{"title":"Sequence-function space of radical SAM cyclophane synthases reveal conserved active site residues that influence substrate specificity.","authors":"Chin-Soon Phan, Brandon I Morinaka","doi":"10.1039/d4cb00227j","DOIUrl":null,"url":null,"abstract":"<p><p>Radical SAM cyclophane synthases catalyze C-C, C-N, and C-O crosslinking reactions in the biosynthesis of bioactive peptide natural products. Here, we studied an uncharacterized rSAM enzyme, HtkB from <i>Pandoraea</i> sp., and found this enzyme to catalyze the formation of a HisC2-to-LysCβ crosslink. We used a combination of ColabFold and mutagenesis studies to show that residues D214 in HtkB and H204 in HaaB (another cyclophane synthase) are important for substrate specificity. Mutation of these residues changes the specificity and lowers substrate recognition on the wild-type motifs. This result opens opportunities to alter the specificity and promiscuity for rSAM peptide modifying enzymes.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":null,"pages":null},"PeriodicalIF":4.2000,"publicationDate":"2024-10-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11499958/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"RSC Chemical Biology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1039/d4cb00227j","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Radical SAM cyclophane synthases catalyze C-C, C-N, and C-O crosslinking reactions in the biosynthesis of bioactive peptide natural products. Here, we studied an uncharacterized rSAM enzyme, HtkB from Pandoraea sp., and found this enzyme to catalyze the formation of a HisC2-to-LysCβ crosslink. We used a combination of ColabFold and mutagenesis studies to show that residues D214 in HtkB and H204 in HaaB (another cyclophane synthase) are important for substrate specificity. Mutation of these residues changes the specificity and lowers substrate recognition on the wild-type motifs. This result opens opportunities to alter the specificity and promiscuity for rSAM peptide modifying enzymes.