Protein-protein interface analysis of the non-ribosomal peptide synthetase peptidyl carrier protein and enzymatic domains

Corpuz, Joshua C.; Sanlley, Javier O.; Burkart, Michael D.

doi:10.1016/j.synbio.2022.02.006

Cited by 22 publications

(22 citation statements)

References 89 publications

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“…While all of the aforementioned methods might lead to the synthesis of the desired NRP product, a significant reduction in product yield is often observed. Meanwhile, the current knowledge of the NRPS systems was mostly based on domain, dual domain and single NRPS module information [ 38 , 39 ]. However, little was known about the interactions between modules in multimodule NRPSs [ 40 ].…”

Section: Discussionmentioning

confidence: 99%

Deletion of COM donor and acceptor domains and the interaction between modules in bacillomycin D produced by Bacillus amyloliquefaciens

Zhang

et al. 2022

Synthetic and Systems Biotechnology

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Section: Discussionmentioning

confidence: 99%

Deletion of COM donor and acceptor domains and the interaction between modules in bacillomycin D produced by Bacillus amyloliquefaciens

Zhang

et al. 2022

Synthetic and Systems Biotechnology

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“…In the crosslinked complexes, both A domains have been resolved in the thiolation conformation (Figure 2a), in which both subdomains are positioned to form a protein interface capable of binding a partner PCP. 5 The A core of PigI and PltF is responsible for binding substrates, ATP and L-proline, whereas the A sub contains residues that are important for catalysis of both the adenylation and thiolation half-reactions. In both crosslinked structures, PigG maintains the canonical 4 α-helix bundle with the PPant attached to Ser36 of PigG and extended into the active site of the A domain, where it is covalently linked to the prolyl-adenosine vinylsulfonamide (Pro-AVSN) chemical probe (Figures 1d and S1).…”

Section: Crystal Structures Of the Pigg−pigi And Pigg−pltfmentioning

confidence: 99%

“…These difficulties can largely be attributed to our limited understanding of the molecular basis of natural product biosynthesis; namely, the sensitive and precise interplay of the key elements and proteins that govern natural product biosynthesis. 5 The work reported herein represents one promising approach toward rational engineering of NRPS pathways.…”

Section: ■ Introductionmentioning

confidence: 99%

Essential Role of Loop Dynamics in Type II NRPS Biomolecular Recognition

et al. 2022

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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.

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“…Ma et al identified a (−)-bornyl diphosphate synthase from Blumea balsamifera and applied it for the biosynthesis of (−)-borneol in yeast [ 1 ]. Corpuz et al reviewed the current progress on protein-protein interface analysis of the non-ribosomal peptide synthetase (NRPS), providing insights for engineering these mega-enzymes [ 2 ]. Similarly, Guzman et al summarized how to use fragment-antigen binding domains as protein crystallization chaperones for structural study of assembly-line polyketide synthases (PKSs), which are of interest to synthesize an unusually broad range of medicinally relevant compounds [ 3 ].…”

Section: Enzyme Characterization and Engineeringmentioning

confidence: 99%