Volume 17 Supplement 7

Selected articles from the International Conference on Intelligent Biology and Medicine (ICIBM) 2015: genomics

Open Access

Genomic data mining reveals a rich repertoire of transport proteins in Streptomyces

BMC Genomics201617(Suppl 7):510

https://doi.org/10.1186/s12864-016-2899-4

Published: 22 August 2016

Abstract

Background

Streptomycetes are soil-dwelling Gram-positive bacteria that are best known as the major producers of antibiotics used in the pharmaceutical industry. The evolution of exceptionally powerful transporter systems in streptomycetes has enabled their adaptation to the complex soil environment.

Results

Our comparative genomic analyses revealed that each of the eleven Streptomyces species examined possesses a rich repertoire of from 761-1258 transport proteins, accounting for 10.2 to 13.7 % of each respective proteome. These transporters can be divided into seven functional classes and 171 transporter families. Among them, the ATP-binding Cassette (ABC) superfamily and the Major Facilitator Superfamily (MFS) represent more than 40 % of all the transport proteins in Streptomyces. They play important roles in both nutrient uptake and substrate secretion, especially in the efflux of drugs and toxicants. The evolutionary flexibility across eleven Streptomyces species is seen in the lineage-specific distribution of transport proteins in two major protein translocation pathways: the general secretory (Sec) pathway and the twin-arginine translocation (Tat) pathway.

Conclusions

Our results present a catalog of transport systems in eleven Streptomyces species. These expansive transport systems are important mediators of the complex processes including nutrient uptake, concentration balance of elements, efflux of drugs and toxins, and the timely and orderly secretion of proteins. A better understanding of transport systems will allow enhanced optimization of production processes for both pharmaceutical and industrial applications of Streptomyces, which are widely used in antibiotic production and heterologous expression of recombinant proteins.

Keywords

Streptomyces Transport proteins Comparative genomics Drug efflux Protein translocation

Background

Streptomyces is a group of soil-dwelling Gram-positive bacteria, which are well known for their ability to produce a broad array of secondary metabolites including antibiotics, antifungals, antiparasitic drugs, anticancer agents, immunosuppressants, and herbicides [1, 2]. They are also ideal systems in biotechnology for heterologous expression of recombinant proteins with simple downstream processing and high yields [3, 4]. In order to survive in the complex soil environment, streptomycetes have evolved exceptionally powerful transport systems [5, 6]. For example, in Streptomyces coelicolor, there are more than 600 predicted transport proteins with a large proportion being the ATP-binding Cassette (ABC) and Major Facilitator Superfamily (MFS) transporters, which have been implicated in the transport of secondary metabolites including antibiotics [7]. In addition to secondary metabolites, streptomycetes also secret to the environment a mass of proteins through the general secretory (Sec) pathway and the twin-arginine translocation (Tat) pathway [810]. These secretory systems are known to facilitate nutrient acquisition. For example, secreted cellulases and chitinases can degrade otherwise insoluble nutrient sources.

Transporters are of critical importance to all living organisms in facilitating metabolism, intercellular communication, biological synthesis and reproduction. They are involved in the uptake of nutrients from the environment, the secretion of metabolites, the efflux of drugs and toxins, the maintenance of ion concentration gradient across membranes, the secretion of macromolecules, such as sugars, lipids, proteins and nucleic acids, signaling molecules, the translocation of membrane proteins, and so on [11]. A Transporter Classification (TC) system has been developed by the Saier group [11, 12]. To date, more than 10,000 non-redundant transport proteins comprising about 750 families are collected in their Transporter Classification Database (TCDB) [13]. These families are divided among seven major classes: Channels/Pores (Class 1), Electrochemical Potential-driven Transporters (Class 2), Primary Active Transporters (Class 3), Group Translocators (Class 4), Transmembrane Electron Carriers (class 5), Accessory Factors Involved in Transport (Class 8), and Incompletely Characterized Transport Systems (Class 9). This classification system has been applied to in-depth studies of transporters in a number of microbial genomes [1417], and is being adopted in this study for Streptomyces.

The availability of genomes from closely related Streptomyces species enables comprehensive analysis of the transport protein families in Streptomyces. In this study, we report a catalog and comparative genomic analysis of transporters in eleven Streptomyces species with complete genome sequences and annotations, including S. coelicolor (SCO), S. avermitilis (SAV), S. bingchenggensis (SBI), S. cattleya (SCAT), S. flavogriseus (SFLA), S. griseus (SGR), S. hygroscopicus (SHJG), S. scabiei (SCAB), S. sp. SirexAA-E (SACTE), S. venezuelae (SVEN) and S. violaceusniger (STRVI) [7, 1824]. We identified and classified these Streptomyces transporters, using the nomenclature in the TCDB. The class, transmembrane topology and substrate specificity of these transporters are investigated in detail. An improved understanding of Streptomyces transporters will bring new insights into the mechanisms underlying the unique and powerful secretion systems of secondary metabolites and proteins in this group of bacteria of enormous economic and biomedical significance.

Results and discussion

Abundant transporters are present in eleven Streptomyces genomes

Strong material intake and secretion capacity powered by transport systems is an adaptive attribute of soil-dwelling bacteria [1]. We used the coding sequences from eleven Streptomyces genomes to query the TCDB [13, 25] using BLASTP and identified 761-1258 transporters in these eleven genomes, which accounted for 10.2 to 13.7 % of each respective proteome (Table 1 and Additional file 1). S. bingchenggensis, which has the largest genome, and the largest number of protein-coding genes, has the largest number of transporters, whereas S. cattleya contains only 761 transporters, the lowest number and proportion of transporters among the eleven Streptomyces species.
Table 1

Distribution of transporters in eleven Streptomyces genomes

Organisms

Accession ID

Genome size (Mbp)

# ORFs

# Transporters

% Transporters

S. avermitilis

NC_003155 (chr)

9.1

7676

989

12.9 %

NC_004719 (pSAP1)

S. bingchenggensis

NC_016582 (chr)

11.9

10022

1258

12.6 %

S. cattleya

NC_016111(chr)

8.1

7475

761

10.2 %

NC_016113(pSCAT)

S. coelicolor

NC_003888(chr)

9.1

8153

990

12.1 %

NC_003903 (pSCP1)

NC_003904 (pSCP2)

S. flavogriseus

NC_016114 (chr)

7.7

6572

888

13.5 %

NC_016110 (pSFLA01)

NC_016115 (pSFLA02)

S. griseus

NC_010572 (chr)

8.5

7136

975

13.7 %

S. hygroscopicus

NC_017765 (chr)

10.4

9108

999

11.0 %

NC_017766 (pSHJG1)

NC_016972 (pSHJG2)

S. scabiei

NC_013929 (chr)

10.1

8746

1021

11.7 %

S. sp. SirexAA-E

NC_015953 (chr)

7.4

6357

869

13.7 %

S. venezuelae

NC_018750 (chr)

8.2

7453

935

12.5 %

S. violaceusniger

NC_015957 (chr)

11.0

8985

989

11.0 %

NC_015951(pSTRVI01)

NC_015952(pSTRVI02)

Streptomyces transporters show diverse transmembrane topology

The capacity of a transporter is often associated with the complexity and topology of its transmembrane region(s) where the major events of substrate uptake or output across the cell membranes take place. Using the TMHMM (TransMembrane prediction using Hidden Markov Models) algorithm [26], we performed the transmembrane topology analysis for Streptomyces transporters to identify the transmembrane segments (TMSs). The number of TMSs ranges from 0 to 24. The largest number of TMSs observed in a transporter in the eleven Streptomyces genomes varies from 16 to 24 (Table 2). Except for intra-/extra-cellular transporters which have no TMS, transporters with 6 and 12 TMSs are predominant. Most transporters with 6 TMSs are ABC transporters (TC 3.A.1), and transporters with 12 TMSs are mainly members of the Major Facilitator Superfamily (MFS) (TC 2.A.1), the Amino Acid-Polyamine-Organocation (APC) superfamily (TC 2.A.3), the Resistance-Nodulation-Cell Division (RND) superfamily (TC 2.A.6) and the ABC superfamily (TC 3.A.1). It is possible that these 12-TMS transporters have arisen from the primordial 6-TMS form via intragenic duplication [27]. Among the transporters with more than 6 TMSs, the transporters with an even number of TMSs are more abundant than those with an odd number of TMSs (Fig. 1). The distribution of TMSs in S. griseus transporters is unique: this bacterium has 53 transporters with 9 TMSs, mostly ABC transporters, accounting for 5.4 % of the total transporters. This proportion is significantly higher than that of the other ten sibling species. On the other hand, S. griseus has the lowest proportion of 12-TMS transporters (7.3 %), most of which are also ABC transporters. These topology patterns suggest that during the evolution of transporters in S. griseus, the “6 + 3” events may be more frequent than the typical “6 + 6” events observed in ten other Streptomyces species [27, 28].
Table 2

Distribution of topological types of transporters in eleven Streptomyces genomes

TMS

SACTE

SAV

SBI

SCAB

SCAT

SCO

SFLA

SGR

SHJG

STRVI

SVEN

0

322

382

482

424

280

344

332

372

392

371

350

1

41

41

55

33

51

47

45

33

28

37

44

2

14

17

18

21

19

19

21

15

16

15

16

3

26

28

32

20

21

26

28

30

22

26

13

4

27

29

31

36

23

32

26

29

36

28

30

5

49

55

62

52

40

58

42

58

56

58

55

6

119

130

201

141

72

135

124

122

116

143

113

7

22

29

23

15

12

24

23

22

20

19

24

8

26

32

35

30

26

34

27

25

35

31

22

9

33

25

36

28

16

35

34

53

30

21

30

10

41

55

62

46

44

54

41

43

48

41

55

11

20

28

42

32

27

30

24

26

40

35

29

12

66

72

109

89

68

89

70

71

97

106

85

13

18

26

24

19

15

20

14

21

25

23

21

14

40

37

43

31

45

38

32

46

35

32

44

15

1

1

1

1

1

1

0

2

0

0

1

16

2

1

1

1

1

1

1

3

2

2

1

17

0

0

1

2

0

2

2

1

1

1

2

18

1

1

0

0

0

0

1

1

0

0

0

19

0

0

0

0

0

0

0

1

0

0

0

24

1

0

0

0

0

1

1

1

0

0

0

Total

869

989

1258

1021

761

990

888

975

999

989

935

Note: SACTE (S. sp. SirexAA-E), SAV (S. avermitilis), SBI (S. bingchenggensis), SCAB (S. scabiei), SCAT (S. cattleya), SCO (S. coelicolor), SFLA (S. flavogriseus), SGR (S. griseus), SHJG (S. hygroscopicus), STRVI (S. violaceusniger), SVEN (S. venezuelae)

Fig. 1

Distribution of transporter topologies in eleven Streptomyces genomes. The abbreviations for species are: S. sp. SirexAA-E (SACTE), S. avermitilis (SAV), S. bingchenggensis (SBI), S. scabiei (SCAB), S. cattleya (SCAT), S. coelicolor (SCO), S. flavogriseus (SFLA), S. griseus (SGR), S. hygroscopicus (SHJG), S. violaceusniger (STRVI), and S. venezuelae (SVEN)

Transporters in eleven Streptomyces genomes can be divided into seven classes and 171 families

The Streptomyces transporters fall into seven classes and 171 transporter families according to the TCDB system (Table 3 and Additional file 2). The distribution of transporters in each species is depicted in Fig. 2.
Table 3

Distribution of Streptomyces transporters in each TC class and subclass

Class

Subclass

SACTE

SAV

SBI

SCAB

SCAT

SCO

SFLA

SGR

SHJG

STRVI

SVEN

1: Channels/Proes

22

31

29

34

22

29

26

28

26

31

30

 

1.A: α-Type Channels

18

24

20

25

15

22

21

20

21

24

21

 

1.B: β-Barrel Porins

3

6

7

8

6

6

4

5

4

6

6

 

1.C: Pore-Forming Toxins (Proteins and Peptides)

1

1

1

1

1

1

1

3

1

1

3

 

1.I: Membrane-bounded channels

0

0

1

0

0

0

0

0

0

0

0

2: Electrochemical Potential-driven Transporters

212

266

330

251

239

274

217

242

305

271

269

 

2.A: Porters (uniporters, symporters, antiporters)

212

266

328

251

239

274

217

242

305

271

269

 

2.C: Ion-gradient-driven energizers

0

0

2

0

0

0

0

0

0

0

0

3: Primary Active Transporters

500

544

705

553

365

552

498

555

489

528

494

 

3.A: P-P-bond-hydrolysis-driven transporters

455

492

656

505

304

497

451

508

433

476

449

 

3.B: Decarboxylation-driven transporters

6

6

5

6

10

7

6

6

6

4

6

 

3.D: Oxidoreduction-driven transporters

39

46

43

42

51

48

41

41

50

47

39

 

3.E: Light absorption-driven transporters

0

0

1

0

0

0

0

0

0

1

0

4: Group Translocators

27

46

62

54

35

30

36

40

46

37

43

 

4.A: Phosphotransfer-driven group translocators

5

7

4

5

2

8

8

6

5

7

6

 

4.B: Nicotinamide ribonucleoside uptake transporters

1

1

0

1

1

1

3

3

1

0

3

 

4.C: Acyl CoA ligase-coupled transporters

21

38

58

48

32

21

25

31

40

30

34

5: Transmembrane Electron Carriers

12

13

21

19

18

26

15

13

20

16

16

 

5.A: Transmembrane 2-electron transfer carriers

12

12

21

18

17

26

14

13

19

15

16

 

5.B: Transmembrane 1-electron transfer carriers

0

1

0

1

1

0

1

0

1

1

0

8: Accessory Factors Involved in Transport

4

4

5

6

5

5

5

6

6

4

4

 

8.A: Auxiliary transport proteins

4

4

5

6

5

5

5

6

6

4

4

9: Incompletely Characterized Transport Systems

60

63

74

67

75

67

68

66

63

69

55

 

9.A: Recognized transporters of unknown biochemical mechanism

27

25

44

27

33

31

32

35

27

33

25

 

9.B: Putative transport proteins

33

38

30

40

42

36

35

31

36

36

30

 

9.C: Functionally characterized transporters lacking identified sequences

0

0

0

0

0

0

1

0

0

0

0

N/A

 

32

22

32

37

2

7

23

25

44

33

24

Total

 

869

989

1258

1021

761

990

888

975

999

989

935

Fig. 2

Distribution of transporter types according to the TC system in eleven Streptomyces genomes. Class 1: Channels/Proes; Class 2: Electrochemical Potential-driven Transporters; Class 3: Primary Active Transporters; Class 4: Group Translocators; Class 5: Transmembrane Electron Carriers; Class 8: Accessory Factors Involved in Transport; Class 9: Incompletely Characterized Transport Systems; N/A: Not assigned

The Primary Active Transporters (Class 3) is the most abundant class of transporters in Streptomyces, which includes 365-705 transporters (representing about 48.0-57.5 % of the total transport machinery). This class of transporters plays important roles in various aspects of bacterial life cycle, especially in the import and export of secondary metabolites, and cation transportation.

Class 2 transporters, the electrochemical potential-driven transporters, are also widely found in Streptomyces. 212-330 transporters in eleven Streptomyces genomes belong to this class, which account for 24.4 %-31.4 % of all the transporters. The porters in this class include uniporters, symporters and antiporters. The most abundant family, MFS, in Class 2 transporters has been implicated in drug efflux. Lineage-specificity is also observed in this class of transporters. For example, S. bingchenggensis possesses two Ion-gradient-driven Energizers (TC 2.C), while the other ten Streptomyces species only have Porters (uniporters, symporters, antiporters) (TC 2.A).

Class 1 transporters are not abundant, but are functionally important for Streptomyces. 22-34 channel/pore transporters are present in these eleven genomes, accounting for 2.3 %-3.2 % of all the transporters. The majority of these channel-type proteins are alpha-type channels (TC 1.A), which have been implicated in stress responses of Gram-positive bacteria, especially responses to osmotic pressure [27]. A small number of proteins belong to β-type porins and a fewer are putative Channel-Forming Toxins (TC 1.C). The membrane-bounded channel (TC 1.I) subclass is rare in Streptomyces; only S. bingchenggensis has a transport protein from this subclass.

Classes 4, 5, and 8 are relatively less abundant. About 3.0 %-5.3 % of all the transport proteins are Class 4 transporters. Two major subclasses observed in Class 4 are the PTS Glucose-Glucoside (Glc) family (4.A.1) and the Fatty Acid Transporter (FAT) family (4.C.1), which are responsible for the transport of glucoses-glucosides and fatty acids, respectively. Notably, S. cattleya, which has the smallest repertoire of transporters among the eleven Streptomyces, does not seem to contain any Glc transporters; it remains unknown if it uses an alternative system. Only 12-21 members of the Class 4 transporters, the Transmembrane Electron Carriers, are found in Streptomyces. Two subclasses are present, including the Prokaryotic Molybdopterin-containing Oxidoreductase (PMO) family (TC 5.A.3) and the Prokaryotic Succinate Dehydrogenase (SDH) family (TC 5.A.4), which transfer electrons mainly by redox reactions. Class 8, the Accessory Factors Involved in Transport, is the least abundant transporter class (0.4 %-0.7 %) in Streptomyces.

A significant number (60-75) of transporters in Streptomyces can be grouped into Class 9, an incompletely characterized class. While their exact physiological roles are yet to be elucidated, they might be involved in the transport of ions, implicated by their sequence similarities with the members of the HlyC/CorC (HCC) family (TC 9.A.40), and the Tripartite Zn2+ Transporter (TZT) family (TC 9.B.10).

Examples of important transporter families

Many of the 171 transporter families are involved in the transfer of ions, saccharides, amino acids, polypeptides, proteins, drugs, toxins and other compounds. The two most abundant and perhaps also the most important families are in the ABC (TC 3.A.1) and MFS (TC 2.A.1) superfamilies. They are responsible for the secretion of a wide array of antibiotics in Streptomyces [29, 30].

The ABC transporters

32.7 %-47.5 % (249-597) of all the transport proteins in the eleven Streptomyces genomes are members of ABC superfamily. ABC transporters are characterized by a conserved ATP hydrolyzing domain for energy provision, pore-forming membrane-integrated domain(s), and a substrate-binding domain [31, 32]. The ABC transport system is composed of the intake system and the efflux system.

The 30 intake families (TC 3.A.1-3.A.33) that we identified in the Streptomyces genomes are specialized in the uptake of diverse nutrient substances. This intake system includes families of Carbohydrate Uptake Transporters (TC 3.A.1.1, 3.A.1.2) that transport saccharides, Polar Amino Acid Uptake Transporters and Hydrophobic Amino Acid Uptake Transporters (TC 3.A.1.3, 3.A.1.4) that transfer amino acids, Polyamine/Opine/ Phosphonate Uptake Transporters and Quaternary Amine Uptake Transporters (TC 3.A.1.11, 3.A.1.12) that transfer amine substances, Iron Chelate Uptake Transporters and Manganese/Zinc/Iron Chelate Uptake Transporters (TC 3.A.1.14, 3.A.1.15) that transfer metal ions.

Unlike the intake system, the 35 Streptomyces efflux families are involved in the transport of macromolecular substances. These transporters are believed to be essential for Streptomyces due to their roles in drug efflux and protein secretion. The drug efflux system regulates various aspects of the response to drug compounds mediated by Drug Exporters (TC 3.A.1.105, 3.A.1.117, 3.A.1.119, 3.A.1.135), Drug Resistance ATPases (TC 3.A.1.120, 3.A.1.121), Macrolide Exporters (TC 3.A.1.122), β-Exotoxin I Exporters (TC 3.A.1.126), Multidrug Resistance Exporters (TC 3.A.201) and Pleiotropic Drug Resistance transporters (TC 3.A.1.205). Potent protein transport in Streptomyces is regulated by Protein/Peptide Exporters (TC 3.A.1.109, 110, 111, 112, 123, 124, 134), Lipoprotein Translocases (TC 3.A.1.125), AmfS Peptide Exporters (TC 3.A.1.127), and SkfA Peptide Exporters (TC 3.A.1.128).

The MFS transporters

Unlike the ABC transporters, the MFS transporters are driven by an electrochemical potential formed by ion concentration gradients across the cytomembrane [30]. There are 90-169 (10.1 %- 15.0 %) MFS transporters in eleven Streptomyces genomes. Streptomyces possesses 39 subfamilies of MFS transporters, including 20 intake systems, 13 efflux systems and 6 systems whose transport direction is unknown. The substances transported by the intake systems are mainly saccharides and organic acids.

One of the most important roles of the MFS transporters is drug efflux [30]. Diverse subfamilies of drug efflux MFS transporters are present in Streptomyces, with varying mechanisms of action, including Drug:H+ Antiporters (TC 2.A.1.2, 2.A.1.3, 2.A.1.21), Aromatic Compound/Drug Exporters (TC 2.A.1.32), Fosmidomycin Resistance transporters (TC 2.A.1.35), Acriflavin-sensitivity transporters (TC 2.A.1.36), and Microcin C51 Immunity Proteins (TC 2.A.1.61), to name a few.

The wide distribution of substrates for Streptomyces transporters

The capacity of the complex and powerful transporter system in Streptomyces is evidenced by the broad scope of the substrates being transported. Figure 3a shows the distribution of transporters that transport different type of substrates in Streptomyces, including carbon sources, drugs, toxicants, electrons, inorganic molecules, macromolecules, amino acids and derivatives, nucleotides and derivatives, vitamins, and accessory factors. The carbon source transporters are the most abundant, with their proportion of all the transport proteins ranging from 21.7 to 31.6 % in eleven genomes. Notably, the substrates of an average of 6.4 % of the transporters in Streptomyces genomes examined cannot be determined based on genomic analysis, and await advanced structural and biochemical characterization.
Fig. 3

a Distribution of substrate types and (b) predicted polar characteristics: bidirectional transport, uptake or export in eleven Streptomyces genomes

Streptomyces transporters can be divided into three classes, uptake, efflux and bidirectional, according to the direction of the substrates transported (Fig. 3b). Among the transporters of the eleven Streptomyces genomes, on average 46.5 % are involved in the uptake of substrates, 35.8 % are involved in the efflux of substrates, and 11.0 % are in charge of the bidirectional transport of substrates. The direction of 6.7 % of these proteins remains undetermined.

Streptomyces have lineage-specific protein secretion systems

Streptomyces have two major lineage-specific protein transport systems, the Tat system (TC 2.A.64) and the Sec system (TC 3.A.5) [8, 9]. The Tat system was shown to be related to the pathogenicity of pathogenic bacteria [33]. In S. scabies, the transporters in the Tat pathway secrete several toxicity-associated proteins [34]. While the key component proteins of the Tat system, TatA, TatB and TatC, are present in all eleven Streptomyces genomes we looked at, lineage-specificity is clearly shown with respect to the copy number variation of these genes (Table 4). Only one copy of the tatB and tatC genes is present in nine Streptomyces genomes; S. flavogriseus has two copies of the tatB genes and S. hygroscopicus has two copies of the tatC genes. The copy number of the tatA gene ranges from one to three in eleven genomes (Table 4). Phylogenetic analysis shows that the multiple copies of the tatA genes may have different evolutionary origins and can be divided into three independent clades, namely tatA1, tatA2 and tatA3 (Fig. 4a). The tatA paralogous genes in the majority of the Streptomyces genomes belong to different clades. Notably, all the three tatA paralogous genes in S. cattleya are clustered into the tatA3 clade, indicative of recent gene duplication events.
Table 4

The Tat translocation system in Streptomyces (TC 2.A.64)

Species

tatA1

tatA2

tatA3

tatB1

tatB2

tatC1

tatC2

SACTE

SACTE_1063

SACTE_6092

SACTE_3032

SACTE_4381

 

SACTE_1062

 

SAV

SAV_6692

  

SAV_3114

 

SAV_6693

 

SBI

SBI_08493

  

SBI_04079

 

SBI_08494

 

SCAB

SCAB_73591

  

SCAB_31121

 

SCAB_73601

 

SCAT

SCAT_3206

SCAT_2668

SCAT_4914

SCAT_4007

 

SCAT_5184

 

SCO

SCO1633

SCO3768

 

SCO5150

 

SCO1632

 

SFLA

Sfla_5203

Sfla_0514

Sfla_5510

Sfla_5507

Sfla_2146

Sfla_5204

 

SGR

SGR_5870

SGR_6484

SGR_340

SGR_2375

 

SGR_5871

 

SHJG

SHJG_2368

SHJG_3070

SHJG_0499

SHJG_6250

 

SHJG_2367

SHJG_3069

STRVI

Strvi_6639

Strvi_3352

 

Strvi_1468

 

Strvi_6638

 

SVEN

SVEN_1225

  

SVEN_4796

 

SVEN_1224

 
Fig. 4

a Phylogenetic tree of the TatA system. b Phylogenetic tree of the SecD/SecF (b) system in eleven Streptomyces genomes. The trees were constructed using the neighbor-joining method by MEGA6 [43]. The Maximum Parsimony and Maximum Likelihood methods gave virtually the same topology (data not shown)

Similarly, the Sec system is also species-specific. This system includes SecA, SecY, SecE, SecG, SecD, SecF, YajC, FtsY, etc. [35], all of which are highly conserved in Streptomyces (Table 5). There is only one copy of the secE, secG, secD, secF, yajC and ftsY genes in each of the eleven Streptomyces genomes. Interestingly, there is a second set of secA2/secY2 genes in several species, which may be involved in the secretion of proteins with specific functions, for example, the secretion of toxic proteins [36]. In S. avermitilis, for instance, there are two copies of the secA genes, and S. venezuelae has two copies of the secY genes.
Table 5

The Sec translocation system in Streptomyces (TC 3.A.5)

Species

secA1

secA2

secY

secY2

secE

secG

SACTE

SACTE_2472

 

SACTE_3988

 

SACTE_3949

SACTE_1366

SAV

SAV_5071

SAV_2565

SAV_4312

 

SAV_4908

SAV_6299

SBI

SBI_06502

 

SBI_06209

 

SBI_06158

SBI_08032

SCAB

SCAB_55371

 

SCAB_36741

 

SCAB_37261

SCAB_69731

SCAT

SCAT_2009

 

SCAT_3612

 

SCAT_3559

SCAT_1102

SCO

SCO3005

 

SCO4722

 

SCO4646

SCO1944

SFLA

Sfla_3902

 

Sfla_2503

 

Sfla_2541

Sfla_4882

SGR

SGR_4531

 

SGR_2814

 

SGR_2876

SGR_5576

SHJG

SHJG_4468

 

SHJG_5817

 

SHJG_5775

SHJG_3400

STRVI

Strvi_8396

 

Strvi_0893

 

Strvi_0854

Strvi_7031

SVEN

SVEN_2748

 

SVEN_4399

SVEN_0354

SVEN_4338

SVEN_1573

Species

secD

secF

secDF

yajC

ftsY

SACTE

SACTE_0919

SACTE_0918

SACTE_5723

SACTE_0920

SACTE_4801

SAV

SAV_6837

SAV_6838

 

SAV_6836

SAV_2654

SBI

SBI_02394

SBI_02393

 

SBI_02395

SBI_03477

SCAB

SCAB_74911

SCAB_74921

SCAB_6041

SCAB_74901

SCAB_26291

SCAT

SCAT_5307

SCAT_5308

 

SCAT_5306

SCAT_4417

SCO

SCO1516

SCO1515

SCO6160

SCO1517

SCO5580

SFLA

Sfla_5348

Sfla_5349

Sfla_0862

Sfla_5347

Sfla_1718

SGR

SGR_6019

SGR_6020

SGR_1134

SGR_6018

SGR_1898

SHJG

SHJG_2940

SHJG_2939

SHJG_8531

SHJG_2941

SHJG_6701

STRVI

Strvi_3032

Strvi_3033

 

Strvi_3031

Strvi_1937

SVEN

SVEN_1116

SVEN_1115

SVEN_0190

SVEN_1117

SVEN_5276

The evolutionary pattern in the secD and the secF genes is particularly interesting (Fig. 4b). In bacteria, these genes encode accessory factors in the Sec pathway that can accelerate the translocation of protein substrates. There are two forms of the secD and secF genes: in the first form, these two genes are adjacent but separate, while in the second form, the two genes are fused into a single secDF gene. The fused secDF is present in seven Streptomyces genomes. Unlike most bacteria that have one of the two forms, the majority of Streptomyces species have both the separated form and the fused form [37]. The acquisition of a second copy may confer a selective advantage to Streptomyces by enhancing the capacity and the effectiveness of protein transport.

Conclusions

Comparative genomic analyses of eleven Streptomyces genomes revealed an abundant repertoire of 761-1258 transporters, belonging to seven transporter classes and 171 transporter families. The powerful transport systems in Streptomyces play critical roles in drug efflux, protein secretion and stress response. A better understanding of transport systems will allow enhanced optimization of production processes for both pharmaceutical and industrial applications of Streptomyces.

Methods

Data

The completed whole genome data of the eleven Streptomyces species (Table 1), including amino acid sequences and functional annotations of all the proteins were downloaded from the NCBI database (http://www.ncbi.nlm.nih.gov/genome/browse/). The transporter classification and amino acid sequences of all classified transporters were downloaded from the TCDB database (http://www.tcdb.org/) [13]. We also collected data from the TransporterDB database [38] (http://www.membranetransport.org/) which included the transporter classification data of S. coelicolor and S. avermitilis, and from the Transporter Inference Parser database [39] (http://biocyc.org/), which identified transporter according to their function annotation and included the relevant data of S. coelicolor, S. avermitilis, S. griseus and S. scabies.

Identification and classification of transporters

The BLASTP search of all the proteins in eleven Streptomyces species versus all the transport proteins in TCDB database was conducted to identify transporters in Streptomyces that are homologs to known and predicted transporters in the TCDB [13, 25]. The threshold for homologous genes was set as follow: E-value ≤ 10-5, similarity ≥ 50 %, and the sequence coverage ≥ 30 %. We classified a Streptomyces transporter based on its homologous gene with known function in the TCDB that had the lowest expected value, the highest similarity score and the highest coverage. The classification of Streptomyces transporters in the TransporterDB and the Transporter Inference Parser, the annotations and the conserved domain information helped to filter false negative and false positive predictions. The Pfam search program based on the Hidden Markov Models (HMMs) (http://pfam.xfam.org/) [40] was used to identify conserved structure domains of Streptomyces transporters, with Pfam GA as the threshold. TMHMM (http://www.cbs.dtu.dk/services/TMHMM/) [26] was used to analyze the transmembrane structures and the number of putative TMSs of Streptomyces transporters.

On the basis of the degree of similarities with known or predicted transporters in the TCDB, as well as the conserved domains and the number and location of TMSs, we further classified the Streptomyces transporters into families and subfamilies of homologous transporters according to the TC system [13]. The TC number generally has five components: V.W.X.Y.Z, representing the transporter class, subclass, family, subfamily and the substrate or range of substrates transported [11, 12]. Most Streptomyces transporters were classified at the transporter family level. The transporters in superfamilies such as ABC and MFS were classified at the subfamily level.

The substrate and transport direction of each Streptomyces transporter was predicted based on homology to functionally characterized transporters in the TCDB. Classification of a putative transporter into a family or subfamily according to the TC system allows for the prediction of substrate types and transport direction with confidence [13, 17, 41].

Phylogenetic analysis of transport protein families

Multiple sequence alignments were obtained using Clustal X 2.1 [42]. Phylogenetic trees were reconstructed using MEGA6 with neighbor-joining (NJ), maximum parsimony (MP) and maximum likelihood (ML) methods [43].

Declarations

Acknowledgements

We thank the Computational Biology Initiative at UTSA for providing computational support. This work was supported by grants from the National Natural Science Foundation of China (31501021) and the Zhejiang Provincial Natural Sciences Foundation of China (LY15C060001) to ZZ, grants from the National Basic Research Program of China (973 Program, 2012CB721005) and the National Natural Science Foundation of China (30870033) to YQL, grants from the National Institutes of Health (GM100806, AI080579, and GM081068) to YW. ZZ was also supported by a government scholarship from the China Scholarship Council. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Declarations

Publication charges for this article have been funded by the National Natural Science Foundation of China (31501021) to ZZ.

This article has been published as part of BMC Genomics Volume 17 Supplement 7, 2016: Selected articles from the International Conference on Intelligent Biology and Medicine (ICIBM) 2015: genomics. The full contents of the supplement are available online at http://bmcgenomics.biomedcentral.com/articles/supplements/volume-17-supplement-7.

Availability of data and materials

The datasets supporting the conclusions of this article are included within the article and its additional files.

Authors’ contributions

YW, YQL and ZZ conceived and designed the study. ZZ, NS, SW and YW performed data analysis. YW and ZZ drafted the manuscript. All authors read and approved the final manuscript.

Competing interests

The authors declare that they have no competing interests.

Consent for publication

Not applicable.

Ethics approval and consent to participate

Not applicable.

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

(1)
College of Pharmaceutical Sciences, Zhejiang University
(2)
Zhejiang Provincial Key Laboratory of Microbial Biochemistry and Metabolism Engineering, Zhejiang University
(3)
Department of Biology and South Texas Center for Emerging Infectious Diseases, University of Texas at San Antonio

References

  1. Hopwood DA. Streptomyces in Nature and Medicine: The Antibiotic Makers. New York: Oxford University Press; 2007.Google Scholar
  2. Garrity GM, Lilburn TG, Cole JR, Harrison SH, Euzéby J., B.J. T. Part 10 - The Bacteria: Phylum “Actinobacteria”: Class Actinobacteria. In: Taxonomic Outline of the Bacteria and Archaea. 2007: Release 7.7.: 399-539.Google Scholar
  3. Anne J, Maldonado B, Van Impe J, Van Mellaert L, Bernaerts K. Recombinant protein production and streptomycetes. J Biotechnol. 2012;158(4):159–67.View ArticlePubMedGoogle Scholar
  4. Li YD, Zhou Z, Lv LX, Hou XP, Li YQ. New approach to achieve high-level secretory expression of heterologous proteins by using Tat signal peptide. Protein Pept Lett. 2009;16(6):706–10.View ArticlePubMedGoogle Scholar
  5. Zhou Z, Gu J, Li YQ, Wang Y. Genome plasticity and systems evolution in Streptomyces. BMC Bioinformatics. 2012;13 Suppl 10:S8.View ArticlePubMedPubMed CentralGoogle Scholar
  6. Zhou Z, Gu J, Du YL, Li YQ, Wang Y. The -omics era- toward a systems-level understanding of streptomyces. Curr Genomics. 2011;12(6):404–16.View ArticlePubMedPubMed CentralGoogle Scholar
  7. Bentley SD, Chater KF, Cerdeno-Tarraga AM, Challis GL, Thomson NR, James KD, Harris DE, Quail MA, Kieser H, Harper D, et al. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature. 2002;417(6885):141–7.View ArticlePubMedGoogle Scholar
  8. Chater KF, Biro S, Lee KJ, Palmer T, Schrempf H. The complex extracellular biology of Streptomyces. FEMS Microbiol Rev. 2010;34(2):171–98.View ArticlePubMedGoogle Scholar
  9. Widdick DA, Dilks K, Chandra G, Bottrill A, Naldrett M, Pohlschroder M, Palmer T. The twin-arginine translocation pathway is a major route of protein export in Streptomyces coelicolor. Proc Natl Acad Sci U S A. 2006;103(47):17927–32.View ArticlePubMedPubMed CentralGoogle Scholar
  10. Morosoli R, Shareck F, Kluepfel D. Protein secretion in streptomycetes. FEMS Microbiol Lett. 1997;146(2):167–74.View ArticlePubMedGoogle Scholar
  11. Busch W, Saier Jr MH. The transporter classification (TC) system, 2002. Crit Rev Biochem Mol Biol. 2002;37(5):287–337.View ArticlePubMedGoogle Scholar
  12. Busch W, Saier Jr MH. The IUBMB-endorsed transporter classification system. Mol Biotechnol. 2004;27(3):253–62.View ArticlePubMedGoogle Scholar
  13. Saier Jr MH, Reddy VS, Tamang DG, Vastermark A. The transporter classification database. Nucleic Acids Res. 2014;42(Database issue):D251–258.View ArticlePubMedGoogle Scholar
  14. Paulsen IT, Nguyen L, Sliwinski MK, Rabus R, Saier Jr MH. Microbial genome analyses: comparative transport capabilities in eighteen prokaryotes. J Mol Biol. 2000;301(1):75–100.View ArticlePubMedGoogle Scholar
  15. Kumar U, Saier Jr MH. Comparative genomic analysis of integral membrane transport proteins in ciliates. J Eukaryot Microbiol. 2015;62(2):167–87.View ArticlePubMedGoogle Scholar
  16. Paparoditis P, Vastermark A, Le AJ, Fuerst JA, Saier Jr MH. Bioinformatic analyses of integral membrane transport proteins encoded within the genome of the planctomycetes species, Rhodopirellula baltica. Biochim Biophys Acta. 2014;1838(1 Pt B):193–215.View ArticlePubMedGoogle Scholar
  17. Getsin I, Nalbandian GH, Yee DC, Vastermark A, Paparoditis PC, Reddy VS, Saier Jr MH. Comparative genomics of transport proteins in developmental bacteria: Myxococcus xanthus and Streptomyces coelicolor. BMC Microbiol. 2013;13:279.View ArticlePubMedPubMed CentralGoogle Scholar
  18. Barbe V, Bouzon M, Mangenot S, Badet B, Poulain J, Segurens B, Vallenet D, Marliere P, Weissenbach J. Complete genome sequence of Streptomyces cattleya NRRL 8057, a producer of antibiotics and fluorometabolites. J Bacteriol. 2011;193(18):5055–6.View ArticlePubMedPubMed CentralGoogle Scholar
  19. Wang XJ, Yan YJ, Zhang B, An J, Wang JJ, Tian J, Jiang L, Chen YH, Huang SX, Yin M, et al. Genome sequence of the milbemycin-producing bacterium streptomyces bingchenggensis. J Bacteriol. 2010;192(17):4526–7.View ArticlePubMed CentralGoogle Scholar
  20. Ohnishi Y, Ishikawa J, Hara H, Suzuki H, Ikenoya M, Ikeda H, Yamashita A, Hattori M, Horinouchi S. Genome sequence of the streptomycin-producing microorganism streptomyces griseus IFO 13350. J Bacteriol. 2008;190(11):4050–60.View ArticlePubMedPubMed CentralGoogle Scholar
  21. Ikeda H, Ishikawa J, Hanamoto A, Shinose M, Kikuchi H, Shiba T, Sakaki Y, Hattori M, Ōmura S. Complete genome sequence and comparative analysis of the industrial microorganism Streptomyces avermitilis. Nat Biotechnol. 2003;21(5):526–31.View ArticlePubMedGoogle Scholar
  22. Bignell DRD, Seipke RF, Huguet-Tapia JC, Chambers AH, Parry RJ, Loria R. Streptomyces scabies 87-22 contains a coronafacic acid-like biosynthetic cluster that contributes to plant-microbe interactions. Mol Plant Microbe Interact. 2010;23(2):161–75.View ArticlePubMedGoogle Scholar
  23. Wu H, Qu S, Lu CY, Zheng HJ, Zhou XF, Bai LQ, Deng ZX. Genomic and transcriptomic insights into the thermo-regulated biosynthesis of validamycin in Streptomyces hygroscopicus 5008. BMC Genomics. 2012;13:337.View ArticlePubMedPubMed CentralGoogle Scholar
  24. Yaxley AM. Study of the complete genome sequence of Streptomyces scabies (or scabiei) 87.22. University of Warwick, Coventry, UK; 2009.Google Scholar
  25. Saier Jr MH, Tran CV, Barabote RD. TCDB: the Transporter Classification Database for membrane transport protein analyses and information. Nucleic Acids Res. 2006;34(Database issue):D181–186.View ArticlePubMedGoogle Scholar
  26. Krogh A, Larsson B, von Heijne G, Sonnhammer EL. Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. J Mol Biol. 2001;305(3):567–80.View ArticlePubMedGoogle Scholar
  27. Saier Jr MH. Tracing pathways of transport protein evolution. Mol Microbiol. 2003;48(5):1145–56.View ArticlePubMedGoogle Scholar
  28. Lam VH, Lee JH, Silverio A, Chan H, Gomolplitinant KM, Povolotsky TL, Orlova E, Sun EI, Welliver CH, Saier Jr MH. Pathways of transport protein evolution: recent advances. Biol Chem. 2011;392(1-2):5–12.View ArticlePubMedPubMed CentralGoogle Scholar
  29. Mendez C, Salas JA. The role of ABC transporters in antibiotic-producing organisms: drug secretion and resistance mechanisms. Res Microbiol. 2001;152(3-4):341–50.View ArticlePubMedGoogle Scholar
  30. Saidijam M, Benedetti G, Ren Q, Xu Z, Hoyle CJ, Palmer SL, Ward A, Bettaney KE, Szakonyi G, Meuller J, et al. Microbial drug efflux proteins of the major facilitator superfamily. Curr Drug Targets. 2006;7(7):793–811.View ArticlePubMedGoogle Scholar
  31. Tomii K, Kanehisa M. A comparative analysis of ABC transporters in complete microbial genomes. Genome Res. 1998;8(10):1048–59.PubMedGoogle Scholar
  32. Wang B, Dukarevich M, Sun EI, Yen MR, Saier Jr MH. Membrane porters of ATP-binding cassette transport systems are polyphyletic. J Membr Biol. 2009;231(1):1–10.View ArticlePubMedPubMed CentralGoogle Scholar
  33. De Buck E, Lammertyn E, Anne J. The importance of the twin-arginine translocation pathway for bacterial virulence. Trends Microbiol. 2008;16(9):442–53.View ArticlePubMedGoogle Scholar
  34. Joshi MV, Mann SG, Antelmann H, Widdick DA, Fyans JK, Chandra G, Hutchings MI, Toth I, Hecker M, Loria R, et al. The twin arginine protein transport pathway exports multiple virulence proteins in the plant pathogen Streptomyces scabies. Mol Microbiol. 2010;77(1):252–71.View ArticleGoogle Scholar
  35. Driessen AJ, Fekkes P, van der Wolk JP. The Sec system. Curr Opin Microbiol. 1998;1(2):216–22.View ArticlePubMedGoogle Scholar
  36. Rigel NW, Braunstein M. A new twist on an old pathway--accessory secretion systems. Mol Microbiol. 2008;69(2):291–302.View ArticlePubMedPubMed CentralGoogle Scholar
  37. Zhou Z, Li Y, Sun N, Sun Z, Lv L, Wang Y, Shen L, Li YQ. Function and evolution of two forms of SecDF homologs in Streptomyces coelicolor. PLoS One. 2014;9(8):e105237.View ArticlePubMedPubMed CentralGoogle Scholar
  38. Ren Q, Chen K, Paulsen IT. TransportDB: a comprehensive database resource for cytoplasmic membrane transport systems and outer membrane channels. Nucleic Acids Res. 2007;35(Database issue):D274–279.View ArticlePubMedGoogle Scholar
  39. Lee TJ, Paulsen I, Karp P. Annotation-based inference of transporter function. Bioinformatics. 2008;24(13):i259–267.View ArticlePubMedPubMed CentralGoogle Scholar
  40. Finn RD, Bateman A, Clements J, Coggill P, Eberhardt RY, Eddy SR, Heger A, Hetherington K, Holm L, Mistry J, et al. Pfam: the protein families database. Nucleic Acids Res. 2014;42(Database issue):D222–230.View ArticlePubMedGoogle Scholar
  41. Saier Jr MH. A functional-phylogenetic classification system for transmembrane solute transporters. Microbiol Mol Biol Rev. 2000;64(2):354–411.View ArticlePubMedPubMed CentralGoogle Scholar
  42. Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, et al. Clustal W and Clustal X version 2.0. Bioinformatics. 2007;23(21):2947–8.View ArticlePubMedGoogle Scholar
  43. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: Molecular evolutionary genetics analysis version 6.0. Mol Biol Evol. 2013;30(12):2725–9.View ArticlePubMedPubMed CentralGoogle Scholar

Copyright

© The Author(s). 2016

Advertisement