scholarly journals Comparative structure, dynamics and evolution of acyl-carrier proteins from Borrelia burgdorferi, Brucella melitensis and Rickettsia prowazekii

2020 ◽  
Vol 477 (2) ◽  
pp. 491-508 ◽  
Author(s):  
Ravi P. Barnwal ◽  
Mandeep Kaur ◽  
Alec Heckert ◽  
Janeka Gartia ◽  
Gabriele Varani
Keyword(s):  
Fatty Acid ◽  
Borrelia Burgdorferi ◽  
Active Sites ◽  
Pathogenic Bacteria ◽  
Fatty Acid Synthase ◽  
Brucella Melitensis ◽  
Carrier Proteins ◽  
Acyl Carrier Proteins ◽  
Rickettsia Prowazekii ◽  
Structure Dynamics

Acyl carrier proteins (ACPs) are small helical proteins found in all kingdoms of life, primarily involved in fatty acid and polyketide biosynthesis. In eukaryotes, ACPs are part of the fatty acid synthase (FAS) complex, where they act as flexible tethers for the growing lipid chain, enabling access to the distinct active sites in FAS. In the type II synthesis systems found in bacteria and plastids, these proteins exist as monomers and perform various processes, from being a donor for synthesis of various products such as endotoxins, to supplying acyl chains for lipid A and lipoic acid FAS (quorum sensing), but also as signaling molecules, in bioluminescence and activation of toxins. The essential and diverse nature of their functions makes ACP an attractive target for antimicrobial drug discovery. Here, we report the structure, dynamics and evolution of ACPs from three human pathogens: Borrelia burgdorferi, Brucella melitensis and Rickettsia prowazekii, which could facilitate the discovery of new inhibitors of ACP function in pathogenic bacteria.

scholarly journals Structure and Dynamic Basis of Molecular Recognition Between Acyltransferase and Carrier Protein in E. coli Fatty Acid Synthesis

2020 ◽  
Author(s):  
Laetitia E. Misson ◽  
Jeffrey T. Mindrebo ◽  
Tony D. Davis ◽  
Ashay Patel ◽  
J. Andrew McCammon ◽  
...  
Keyword(s):  
Molecular Dynamics ◽  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Polyketide Synthase ◽  
De Novo ◽  
Md Simulations ◽  
Type Ii ◽  
Carrier Proteins ◽  
Acyl Carrier Proteins ◽  
Binding Interface

AbstractFatty acid synthases (FASs) and polyketide synthases (PKSs) iteratively elongate and often reduce two-carbon ketide units in de novo fatty acid and polyketide biosynthesis. Cycles of chain extensions in FAS and PKS are initiated by an acyltransferase (AT), which loads monomer units onto acyl carrier proteins (ACPs), small, flexible proteins that shuttle covalently linked intermediates between catalytic partners. Formation of productive ACP-AT interactions is required for catalysis and specificity within primary and secondary FAS and PKS pathways. Here, we use the Escherichia coli FAS AT, FabD, and its cognate ACP, AcpP, to interrogate type II FAS ACP-AT interactions. We utilize a covalent crosslinking probe to trap transient interactions between AcpP and FabD to elucidate the first x-ray crystal structure of a type II ACP-AT complex. Our structural data are supported using a combination of mutational, crosslinking, and kinetic analyses, and long timescale molecular dynamics (MD) simulations. Together, these complementary approaches reveal key catalytic features of FAS ACP-AT interactions. These mechanistic inferences suggest that AcpP adopts multiple, productive conformations at the AT binding interface, allowing the complex to sustain high transacylation rates. Furthermore, MD simulations support rigid body subdomain motions within the FabD structure that may play a key role in AT activity and substrate selectivity.Significance StatementThe essential role of acyltransferases (ATs) in fatty acid synthase (FAS) and polyketide synthase (PKS) pathways, namely the selection and loading of starter and extender units onto acyl carrier proteins (ACPs), relies on catalytically productive ACP-AT interactions. Here, we describe and interrogate the first structure of a type II FAS malonyl-CoA:ACP-transacylase (MAT) in covalent complex with its cognate ACP. We combine structural, mutational, crosslinking and kinetic data with molecular dynamics simulations to describe a highly flexible and robust protein-protein interface, substrate-induced active site reorganization, and key subdomain motions that likely govern FAS function. These findings strengthen a mechanistic understanding of molecular recognitions between ACPs and partner enzymes and provide new insights for engineering AT-dependent biosynthetic pathways.

scholarly journals Engineered chimeras unveil swappable modular features of fatty acid and polyketide synthase acyl carrier proteins

2021 ◽  
Author(s):  
Yae In Cho ◽  
Claire L Armstrong ◽  
Ariana Sulpizio ◽  
Kofi K Acheampong ◽  
Kameron N Banks ◽  
...  
Keyword(s):  
Natural Products ◽  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Polyketide Synthase ◽  
Building Blocks ◽  
Combinatorial Biosynthesis ◽  
Biosynthetic Pathways ◽  
Carrier Proteins ◽  
Acyl Carrier Proteins ◽  
E Coli

The strategic redesign of microbial biosynthetic pathways is a compelling route to access molecules of diverse structure and function in a potentially environmentally sustainable fashion. The promise of this approach hinges on an improved understanding of acyl carrier proteins (ACPs), which serve as central hubs in biosynthetic pathways. These small, flexible proteins mediate the transport of molecular building blocks and intermediates to enzymatic partners that extend and tailor the growing natural products. Past combinatorial biosynthesis efforts have failed due to incompatible ACP-enzyme pairings. Herein we report the design of chimeric ACPs with features of the actinorhodin polyketide synthase ACP (ACT) and of the E. coli fatty acid synthase (FAS) ACP (AcpP). We evaluate the ability of the chimeric ACPs to interact with the E. coli FAS ketosynthase FabF, which represents an interaction essential to building the carbon backbone of the synthase molecular output. Given that AcpP interacts with FabF but ACT does not, we sought to exchange modular features of ACT with AcpP to confer functionality with FabF. The interactions of chimeric ACPs with FabF were interrogated using sedimentation velocity experiments, surface plasmon resonance analyses, mechanism-based crosslinking assays, and molecular dynamics simulations. Results suggest that the residues guiding AcpP-FabF compatibility and ACT-FabF incompatibility may reside in the loop I, α-helix II region. These findings can inform the development of strategic secondary element swaps that expand the enzyme compatibility of ACPs across systems and therefore represent a critical step towards the strategic engineering of unnatural natural products.

Acyl carrier protein: structure–function relationships in a conserved multifunctional protein family

2007 ◽  
Vol 85 (6) ◽  
pp. 649-662 ◽  
Author(s):  
David M. Byers ◽  
Huansheng Gong
Keyword(s):  
Fatty Acid ◽  
Active Sites ◽  
Fatty Acid Synthase ◽  
Acyl Carrier Protein ◽  
Divalent Cations ◽  
Structural Features ◽  
Carrier Protein ◽  
Type I ◽  
Prosthetic Group

Acyl carrier protein (ACP) is a universal and highly conserved carrier of acyl intermediates during fatty acid synthesis. In yeast and mammals, ACP exists as a separate domain within a large multifunctional fatty acid synthase polyprotein (type I FAS), whereas it is a small monomeric protein in bacteria and plastids (type II FAS). Bacterial ACPs are also acyl donors for synthesis of a variety of products, including endotoxin and acylated homoserine lactones involved in quorum sensing; the distinct and essential nature of these processes in growth and pathogenesis make ACP-dependent enzymes attractive antimicrobial drug targets. Additionally, ACP homologues are key components in the production of secondary metabolites such as polyketides and nonribosomal peptides. Many ACPs exhibit characteristic structural features of natively unfolded proteins in vitro, with a dynamic and flexible conformation dominated by 3 parallel α helices that enclose the thioester-linked acyl group attached to a phosphopantetheine prosthetic group. ACP conformation may also be influenced by divalent cations and interaction with partner enzymes through its “recognition” helix II, properties that are key to its ability to alternately sequester acyl groups and deliver them to the active sites of ACP-dependent enzymes. This review highlights recent progress in defining how the structural features of ACP are related to its multiple carrier roles in fatty acid metabolism.

scholarly journals A General Method for Quantification and Discovery of Acyl Groups Attached to Acyl Carrier Proteins in Fatty Acid Metabolism Using LC-MS/MS

2020 ◽  
Vol 32 (4) ◽  
pp. 820-832
Author(s):  
Jeong-Won Nam ◽  
Lauren M. Jenkins ◽  
Jia Li ◽  
Bradley S. Evans ◽  
Jan G. Jaworski ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Carrier Proteins ◽  
Acyl Carrier Proteins ◽  
General Method

scholarly journals Structural and dynamical rationale for fatty acid unsaturation inEscherichia coli

2019 ◽  
Vol 116 (14) ◽  
pp. 6775-6783 ◽  
Author(s):  
Greg J. Dodge ◽  
Ashay Patel ◽  
Kara L. Jaremko ◽  
J. Andrew McCammon ◽  
Janet L. Smith ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Active Sites ◽  
Fatty Acid Biosynthesis ◽  
Binding Pocket ◽  
Cross Linking ◽  
Substrate Selectivity ◽  
Acid Biosynthesis ◽  
Acyl Carrier Proteins ◽  
Dynamics Simulations

Fatty acid biosynthesis in α- and γ-proteobacteria requires two functionally distinct dehydratases, FabA and FabZ. Here, mechanistic cross-linking facilitates the structural characterization of a stable hexameric complex of sixEscherichia coliFabZ dehydratase subunits with six AcpP acyl carrier proteins. The crystal structure sheds light on the divergent substrate selectivity of FabA and FabZ by revealing distinct architectures of the binding pocket. Molecular dynamics simulations demonstrate differential biasing of substrate orientations and conformations within the active sites of FabA and FabZ such that FabZ is preorganized to catalyze only dehydration, while FabA is primed for both dehydration and isomerization.

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