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Assembly line polyketide synthases: mechanistic insights and unsolved problems.

Khosla C, Herschlag D, Cane DE, Walsh CT - Biochemistry (2014)

Bottom Line: Enzymol.Relat.Areas Mol.

View Article: PubMed Central - PubMed

Affiliation: Departments of Chemical Engineering, Chemistry, and Biochemistry, Stanford University , Stanford, California 94305, United States.

ABSTRACT
Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75-106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain-domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology.

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Ribbon diagram representationsof atomic structures of prototypicaldomains and didomains from assembly line polyketide synthases. Infigures showing KR and ER domains, the bound NADPH cofactor is alsoshown. All structures were derived from components of DEBS itself,with the exception of the ER-KR didomain obtained from the spinosynsynthase. For details, see refs (4−9).
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fig2: Ribbon diagram representationsof atomic structures of prototypicaldomains and didomains from assembly line polyketide synthases. Infigures showing KR and ER domains, the bound NADPH cofactor is alsoshown. All structures were derived from components of DEBS itself,with the exception of the ER-KR didomain obtained from the spinosynsynthase. For details, see refs (4−9).

Mentions: A fundamental understandingof the operation and specificity ofany assembly line must be built upon knowledge of its structure. Asshown in Figure 1A, DEBS is composed of sixmultifunctional protein modules, each of which is responsible fora single round of polyketide chain elongation and functional groupmodification. Each module is in turn composed of specific combinationsof catalytic domains that catalyze the individual biochemical stepsof chain elongation and processing. The domain organization of module3 of DEBS is shown in Figure 1B, as is theacyl carrier protein (ACP) domain from the upstream module that suppliesmodule 3 with its substrate and the ketosynthase (KS) domain fromthe downstream module that receives its product. By now, the atomicstructures of one or more prototypical members of every domain familyfound within DEBS have been determined (Figure 2).4−9 In addition to providing snapshots of the components of the biosyntheticassembly line, these structures also allow deeper analysis of thecatalytic chemistry mediated by each domain.


Assembly line polyketide synthases: mechanistic insights and unsolved problems.

Khosla C, Herschlag D, Cane DE, Walsh CT - Biochemistry (2014)

Ribbon diagram representationsof atomic structures of prototypicaldomains and didomains from assembly line polyketide synthases. Infigures showing KR and ER domains, the bound NADPH cofactor is alsoshown. All structures were derived from components of DEBS itself,with the exception of the ER-KR didomain obtained from the spinosynsynthase. For details, see refs (4−9).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4020578&req=5

fig2: Ribbon diagram representationsof atomic structures of prototypicaldomains and didomains from assembly line polyketide synthases. Infigures showing KR and ER domains, the bound NADPH cofactor is alsoshown. All structures were derived from components of DEBS itself,with the exception of the ER-KR didomain obtained from the spinosynsynthase. For details, see refs (4−9).
Mentions: A fundamental understandingof the operation and specificity ofany assembly line must be built upon knowledge of its structure. Asshown in Figure 1A, DEBS is composed of sixmultifunctional protein modules, each of which is responsible fora single round of polyketide chain elongation and functional groupmodification. Each module is in turn composed of specific combinationsof catalytic domains that catalyze the individual biochemical stepsof chain elongation and processing. The domain organization of module3 of DEBS is shown in Figure 1B, as is theacyl carrier protein (ACP) domain from the upstream module that suppliesmodule 3 with its substrate and the ketosynthase (KS) domain fromthe downstream module that receives its product. By now, the atomicstructures of one or more prototypical members of every domain familyfound within DEBS have been determined (Figure 2).4−9 In addition to providing snapshots of the components of the biosyntheticassembly line, these structures also allow deeper analysis of thecatalytic chemistry mediated by each domain.

Bottom Line: Enzymol.Relat.Areas Mol.

View Article: PubMed Central - PubMed

Affiliation: Departments of Chemical Engineering, Chemistry, and Biochemistry, Stanford University , Stanford, California 94305, United States.

ABSTRACT
Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75-106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain-domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology.

Show MeSH