| Structural highlights
Function
PIGI_SERS3 Involved in the biosynthesis of 4-methoxy-2,2'-bipyrrole-5-carbaldehyde (MBC), one of the terminal products involved in the biosynthesis of the red antibiotic prodigiosin (Pig) (PubMed:15853884, PubMed:17002325). Catalyzes the conversion of L-proline to L-prolyl-AMP and the transfer of the L-prolyl group to acyl carrier protein PigG (PubMed:17002325).[1] [2]
Publication Abstract from PubMed
Non-ribosomal peptides play a critical role in the clinic as therapeutic agents. To access more chemically diverse therapeutics, non-ribosomal peptide synthetases (NRPSs) have been targeted for engineering through combinatorial biosynthesis; however, this has been met with limited success in part due to the lack of proper protein-protein interactions between non-cognate proteins. Herein, we report our use of chemical biology to enable X-ray crystallography, molecular dynamics (MD) simulations, and biochemical studies to elucidate binding specificities between peptidyl carrier proteins (PCPs) and adenylation (A) domains. Specifically, we determined X-ray crystal structures of a type II PCP crosslinked to its cognate A domain, PigG and PigI, and of PigG crosslinked to a non-cognate PigI homologue, PltF. The crosslinked PCP-A domain structures possess large protein-protein interfaces that predominantly feature hydrophobic interactions, with specific electrostatic interactions that orient the substrate for active site delivery. MD simulations of the PCP-A domain complexes and unbound PCP structures provide a dynamical evaluation of the transient interactions formed at PCP-A domain interfaces, which confirm the previously hypothesized role of a PCP loop as a crucial recognition element. Finally, we demonstrate that the interfacial interactions at the PCP loop 1 region can be modified to control PCP binding specificity through gain-of-function mutations. This work suggests that loop conformational preferences and dynamism account for improved shape complementary in the PCP-A domain interactions. Ultimately, these studies show how crystallographic, biochemical, and computational methods can be used to rationally re-engineer NRPSs for non-cognate interactions.
Essential Role of Loop Dynamics in Type II NRPS Biomolecular Recognition.,Corpuz JC, Patel A, Davis TD, Podust LM, McCammon JA, Burkart MD ACS Chem Biol. 2022 Oct 21;17(10):2890-2898. doi: 10.1021/acschembio.2c00523. , Epub 2022 Sep 29. PMID:36173802[3]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
- ↑ Williamson NR, Simonsen HT, Ahmed RA, Goldet G, Slater H, Woodley L, Leeper FJ, Salmond GP. Biosynthesis of the red antibiotic, prodigiosin, in Serratia: identification of a novel 2-methyl-3-n-amyl-pyrrole (MAP) assembly pathway, definition of the terminal condensing enzyme, and implications for undecylprodigiosin biosynthesis in Streptomyces. Mol Microbiol. 2005 May;56(4):971-89. PMID:15853884 doi:http://dx.doi.org/10.1111/j.1365-2958.2005.04602.x
- ↑ Garneau-Tsodikova S, Dorrestein PC, Kelleher NL, Walsh CT. Protein assembly line components in prodigiosin biosynthesis: characterization of PigA,G,H,I,J. J Am Chem Soc. 2006 Oct 4;128(39):12600-1. PMID:17002325 doi:http://dx.doi.org/10.1021/ja063611l
- ↑ Corpuz JC, Patel A, Davis TD, Podust LM, McCammon JA, Burkart MD. Essential Role of Loop Dynamics in Type II NRPS Biomolecular Recognition. ACS Chem Biol. 2022 Oct 21;17(10):2890-2898. doi: 10.1021/acschembio.2c00523. , Epub 2022 Sep 29. PMID:36173802 doi:http://dx.doi.org/10.1021/acschembio.2c00523
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