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A systematic computational analysis of biosynthetic gene cluster evolution: lessons for engineering biosynthesis.

Medema MH, Cimermancic P, Sali A, Takano E, Fischbach MA - PLoS Comput. Biol. (2014)

Bottom Line: Attempts to engineer their biosynthetic gene clusters (BGCs) to produce unnatural metabolites with improved properties are often frustrated by the unpredictability and complexity of the enzymes that synthesize these molecules, suggesting that genetic changes within BGCs are limited by specific constraints.Here, by performing a systematic computational analysis of BGC evolution, we derive evidence for three findings that shed light on the ways in which, despite these constraints, nature successfully invents new molecules: 1) BGCs for complex molecules often evolve through the successive merger of smaller sub-clusters, which function as independent evolutionary entities. 2) An important subset of polyketide synthases and nonribosomal peptide synthetases evolve by concerted evolution, which generates sets of sequence-homogenized domains that may hold promise for engineering efforts since they exhibit a high degree of functional interoperability, 3) Individual BGC families evolve in distinct ways, suggesting that design strategies should take into account family-specific functional constraints.These findings suggest novel strategies for using synthetic biology to rationally engineer biosynthetic pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbial Physiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands; Groningen Bioinformatics Centre, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.

ABSTRACT
Bacterial secondary metabolites are widely used as antibiotics, anticancer drugs, insecticides and food additives. Attempts to engineer their biosynthetic gene clusters (BGCs) to produce unnatural metabolites with improved properties are often frustrated by the unpredictability and complexity of the enzymes that synthesize these molecules, suggesting that genetic changes within BGCs are limited by specific constraints. Here, by performing a systematic computational analysis of BGC evolution, we derive evidence for three findings that shed light on the ways in which, despite these constraints, nature successfully invents new molecules: 1) BGCs for complex molecules often evolve through the successive merger of smaller sub-clusters, which function as independent evolutionary entities. 2) An important subset of polyketide synthases and nonribosomal peptide synthetases evolve by concerted evolution, which generates sets of sequence-homogenized domains that may hold promise for engineering efforts since they exhibit a high degree of functional interoperability, 3) Individual BGC families evolve in distinct ways, suggesting that design strategies should take into account family-specific functional constraints. These findings suggest novel strategies for using synthetic biology to rationally engineer biosynthetic pathways.

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Qualitative model for the evolution of NRPS/PKS domains.After modules are duplicated, they may get ‘trapped’ in a cycle in which small sequence divergences are counterbalanced by internal recombinations that drive concerted evolution. Through strong diversifying selection (or sufficient drift), domains may break out of this cycle towards domain sequences that are protected from concerted evolution by functional divergence and subsequent stabilizing selection on the new function, or by reduced internal recombination rates due to larger sequence differences between the domains. The abovementioned sequence divergence may occur through cumulative mutation or through recombination with other gene clusters (or other modules within the same gene cluster).
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pcbi-1004016-g004: Qualitative model for the evolution of NRPS/PKS domains.After modules are duplicated, they may get ‘trapped’ in a cycle in which small sequence divergences are counterbalanced by internal recombinations that drive concerted evolution. Through strong diversifying selection (or sufficient drift), domains may break out of this cycle towards domain sequences that are protected from concerted evolution by functional divergence and subsequent stabilizing selection on the new function, or by reduced internal recombination rates due to larger sequence differences between the domains. The abovementioned sequence divergence may occur through cumulative mutation or through recombination with other gene clusters (or other modules within the same gene cluster).

Mentions: Concerted evolution is not peculiar to the rapamycin family (Figure S8). For the gene clusters encoding the biosynthesis of the mutually closely related macrolides erythromycin [31], oleandomycin [32] and pikromycin [33], BGC-specific branching appeared to occur for both KS and AT domains, similar to the pattern for rapamycin, FK520, meridamycin and pladienolide. However, for the ansamycin antibiotics macbecin [34], geldanamycin [35] and herbimycin [36], and the antifungals pimaricin [37], nystatin [38] and amphotericin [39], BGC-specific branching occurs only for AT domains, and not for KS domains. Finally, corroborating earlier observations [40], domains from the trans-AT PKS gene clusters encoding pederin [41] and psymberin [42] do not show any BGC-specific branching at all. We observed that certain NRPS gene clusters also show signs of concerted evolution: a clear BGC-specific branching pattern pointing to concerted evolution can be seen for the A domains and most of the C domains of the gene clusters encoding the biosynthesis of the closely related calcium-dependent lipopeptides daptomycin [23], A54145 [43] and CDA [44]. However, the glycopeptide gene clusters encoding the biosynthesis of balhimycin [45], teicoplanin [46] and A40926 [47] showed no such pattern at all: almost all domains cluster in groups corresponding to domains in the same positions in the assembly line. Collectively, these observations suggest that concerted evolution is a key mechanism driving the evolution of NRPS and PKS gene sequences, but the extent to which it happens depends on family-specific functional constraints as well as on the presence of other evolutionary forces acting upon a gene cluster. Our qualitative model of PKS/NRPS evolution (Fig. 4), which summarizes the interplay of concerted evolution with other evolutionary mechanisms, is relevant to PKS/NRPS engineering efforts: the highly homologous sets of domains generated by concerted evolution are more likely to be mutually interoperable than domain sets chosen at random, and might therefore be attractive building blocks for synthetic biological engineering of biosynthetic pathways.


A systematic computational analysis of biosynthetic gene cluster evolution: lessons for engineering biosynthesis.

Medema MH, Cimermancic P, Sali A, Takano E, Fischbach MA - PLoS Comput. Biol. (2014)

Qualitative model for the evolution of NRPS/PKS domains.After modules are duplicated, they may get ‘trapped’ in a cycle in which small sequence divergences are counterbalanced by internal recombinations that drive concerted evolution. Through strong diversifying selection (or sufficient drift), domains may break out of this cycle towards domain sequences that are protected from concerted evolution by functional divergence and subsequent stabilizing selection on the new function, or by reduced internal recombination rates due to larger sequence differences between the domains. The abovementioned sequence divergence may occur through cumulative mutation or through recombination with other gene clusters (or other modules within the same gene cluster).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256081&req=5

pcbi-1004016-g004: Qualitative model for the evolution of NRPS/PKS domains.After modules are duplicated, they may get ‘trapped’ in a cycle in which small sequence divergences are counterbalanced by internal recombinations that drive concerted evolution. Through strong diversifying selection (or sufficient drift), domains may break out of this cycle towards domain sequences that are protected from concerted evolution by functional divergence and subsequent stabilizing selection on the new function, or by reduced internal recombination rates due to larger sequence differences between the domains. The abovementioned sequence divergence may occur through cumulative mutation or through recombination with other gene clusters (or other modules within the same gene cluster).
Mentions: Concerted evolution is not peculiar to the rapamycin family (Figure S8). For the gene clusters encoding the biosynthesis of the mutually closely related macrolides erythromycin [31], oleandomycin [32] and pikromycin [33], BGC-specific branching appeared to occur for both KS and AT domains, similar to the pattern for rapamycin, FK520, meridamycin and pladienolide. However, for the ansamycin antibiotics macbecin [34], geldanamycin [35] and herbimycin [36], and the antifungals pimaricin [37], nystatin [38] and amphotericin [39], BGC-specific branching occurs only for AT domains, and not for KS domains. Finally, corroborating earlier observations [40], domains from the trans-AT PKS gene clusters encoding pederin [41] and psymberin [42] do not show any BGC-specific branching at all. We observed that certain NRPS gene clusters also show signs of concerted evolution: a clear BGC-specific branching pattern pointing to concerted evolution can be seen for the A domains and most of the C domains of the gene clusters encoding the biosynthesis of the closely related calcium-dependent lipopeptides daptomycin [23], A54145 [43] and CDA [44]. However, the glycopeptide gene clusters encoding the biosynthesis of balhimycin [45], teicoplanin [46] and A40926 [47] showed no such pattern at all: almost all domains cluster in groups corresponding to domains in the same positions in the assembly line. Collectively, these observations suggest that concerted evolution is a key mechanism driving the evolution of NRPS and PKS gene sequences, but the extent to which it happens depends on family-specific functional constraints as well as on the presence of other evolutionary forces acting upon a gene cluster. Our qualitative model of PKS/NRPS evolution (Fig. 4), which summarizes the interplay of concerted evolution with other evolutionary mechanisms, is relevant to PKS/NRPS engineering efforts: the highly homologous sets of domains generated by concerted evolution are more likely to be mutually interoperable than domain sets chosen at random, and might therefore be attractive building blocks for synthetic biological engineering of biosynthetic pathways.

Bottom Line: Attempts to engineer their biosynthetic gene clusters (BGCs) to produce unnatural metabolites with improved properties are often frustrated by the unpredictability and complexity of the enzymes that synthesize these molecules, suggesting that genetic changes within BGCs are limited by specific constraints.Here, by performing a systematic computational analysis of BGC evolution, we derive evidence for three findings that shed light on the ways in which, despite these constraints, nature successfully invents new molecules: 1) BGCs for complex molecules often evolve through the successive merger of smaller sub-clusters, which function as independent evolutionary entities. 2) An important subset of polyketide synthases and nonribosomal peptide synthetases evolve by concerted evolution, which generates sets of sequence-homogenized domains that may hold promise for engineering efforts since they exhibit a high degree of functional interoperability, 3) Individual BGC families evolve in distinct ways, suggesting that design strategies should take into account family-specific functional constraints.These findings suggest novel strategies for using synthetic biology to rationally engineer biosynthetic pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbial Physiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands; Groningen Bioinformatics Centre, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.

ABSTRACT
Bacterial secondary metabolites are widely used as antibiotics, anticancer drugs, insecticides and food additives. Attempts to engineer their biosynthetic gene clusters (BGCs) to produce unnatural metabolites with improved properties are often frustrated by the unpredictability and complexity of the enzymes that synthesize these molecules, suggesting that genetic changes within BGCs are limited by specific constraints. Here, by performing a systematic computational analysis of BGC evolution, we derive evidence for three findings that shed light on the ways in which, despite these constraints, nature successfully invents new molecules: 1) BGCs for complex molecules often evolve through the successive merger of smaller sub-clusters, which function as independent evolutionary entities. 2) An important subset of polyketide synthases and nonribosomal peptide synthetases evolve by concerted evolution, which generates sets of sequence-homogenized domains that may hold promise for engineering efforts since they exhibit a high degree of functional interoperability, 3) Individual BGC families evolve in distinct ways, suggesting that design strategies should take into account family-specific functional constraints. These findings suggest novel strategies for using synthetic biology to rationally engineer biosynthetic pathways.

Show MeSH