Mechanistic and Structural Insights into a Divergent PLP-Dependent l -Enduracididine Cyclase from a Toxic Cyanobacterium
Year: 2023
Type: article
Abstract: ABSTRACT Cyclic arginine noncanonical amino acids (ncAAs) are found in several actinobacterial peptide natural products with therapeutically useful antibacterial properties. The preparation of ncAAs like enduracididine and capreomycidine currently takes multiple biosynthetic or chemosynthetic steps, thus limiting the commercial availability and applicability of these cyclic guanidine-containing amino acids. We recently discovered and characterized the biosynthetic pathway of guanitoxin, a potent freshwater cya-nobacterial neurotoxin, that contains an arginine-derived cyclic guanidine phosphate within its highly polar structure. The ncAA L-enduracididine is an early intermediate in guanitoxin biosynthesis and is produced by GntC, a unique pyridoxal-5’-phosphate (PLP)-dependent enzyme. GntC catalyzes a cyclodehydration from a stereoselectively γ-hydroxylated L-arginine precursor via a reaction that functionally and mechanistically diverges from previously established actinobacterial cyclic arginine ncAA pathways. Herein, we interrogate L-enduracididine biosynthesis from the cyanobacterium Sphaerospermopsis torques-reginae ITEP-024 using spectroscopic, stable isotope labeling techniques, and X-ray crystal structure-guided site-directed mutagenesis. GntC initially facilitates the reversible deprotonations of the α- and β-positions of its substrate prior to catalyzing an irreversible diastereoselective dehydration and subsequent intramolecular cyclization. The comparison of holo- and substrate bound GntC structures and activity assays on sitespecific mutants further identified amino acid residues that contribute to the overall catalytic mechanism. These interdisciplinary efforts at structurally and functionally characterizing GntC enables an improved understanding of how Nature divergently produces cyclic arginine ncAAs and generates additional tools for their biocatalytic production and downstream biological applications.
Source: ACS Catalysis
Language: English
Subfield: Applied Microbiology and Biotechnology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: hybrid
Estimated APC paid: $5,000
Funders: U.S. Department of Energy, Simons Foundation, University of California, Santa Cruz, Life Sciences Research Foundation, Fundação de Amparo à Pesquisa do Estado de São Paulo, National Institutes of Health, Office of Science, National Institute of General Medical Sciences, National Institute of Environmental Health Sciences, Basic Energy Sciences, Biological and Environmental Research, SLAC National Accelerator Laboratory
Awards: G1229526393, G1286236842, G1286251239, G1337544421, G1451874870, G1540461676, G1767990418, G1804828992, G1831385526, G223002938, G2371548206, G2539837713, G2603595557, G262532246, G303061748, G30714459, G3083819904, G315621713, Baccalaureate Bridge to the Biomedical Sciences Program (ACCESS), G334911014, G3488436650, G3629260272, G382641700, G419881185, G4235786860, G4280418065, G42916606, G4501827968, G4581610328, Biosynthesis and Monitoring of the Cyanobacterial Toxin Anatoxin-a(s), G495301720, G498139845, G5570207115, G6173709932, G6188209572, G650090426, G6550344761, G6652958823, G7283024673, G7787287669, G7912248388, G8229677252, G8382000481, A Synchrotron Radiation Structural Biology Resource, G8925327636, G8973213077, ALS Efficiently Networking Advanced Beam Line Experiments (ALS-ENABLE), G911061179, G969889393
OpenAlex ID: W4383875873