Ternary complexes of isopentenyl phosphate kinase from Thermococcus paralvinellae reveal molecular determinants of non‐natural substrate specificity

• DOI: 10.1002/prot.26674
• Publication Date: 2024-02-09T10:16:36Z
• Authors: Bryce P. Johnson; Prashant S. Mandal; Sara M. Brown; Leonard M. Thomas; Shanteri Singh
• Keywords: Ternary complex; Thermococcus

Abstract Isopentenyl phosphate kinases (IPKs) have recently garnered attention for their central role in biocatalytic “isoprenol pathways,” which seek to reduce the synthesis of isoprenoid precursors two enzymatic steps. Furthermore, natural promiscuity IPKs toward non‐natural alkyl‐monophosphates (alkyl‐Ps) as substrates has hinted at isoprenol pathways' potential access novel isoprenoids with potentially useful activities. However, only a handful IPK crystal structures been solved date, and even fewer these contain bound active site. The current study sought elucidate additional ternary complexes using homolog from Thermococcus paralvinellae ( Tcp IPK). Four such were solved, each different alkyl‐P phosphoryl donor substrate/product adenosine triphosphate (ATP)/adenosine diphosphate (ADP). As expected, quaternary, tertiary, secondary closely resembled those published previously, kinetic analysis substrate highlighted dramatic effects altering core scaffold substrate. Even more interesting, though, was discovery trend correlating position α helices site magnitude an homolog's reaction rate reaction. Overall, highlight importance continued structural better understand optimize activity both substrates.