Deciphering mutational effects on inducible NO synthase conformational dynamics via quantitative cross-linking mass spectrometry and AlphaFold2 subsampling.

ABSTRACT: Mammalian nitric oxide synthases (NOSs) are flavo-hemoproteins that rely on dynamic interdomain interactions for activity. Calmodulin (CaM) facilitates specific, interdomain FMN-heme interactions that enable inter-subunit FMN-heme electron transfer essential for nitric oxide biosynthesis. Our quantitative cross-linking mass spectrometry (qXL MS) results demonstrate that the abundance of intersubunit cross-links correlates with CaM-induced formation of the docked FMN/heme complex in rat neuronal NOS [Jiang et al., Biochemistry, 2024, 63, 1395-1411]. Here, we extend this methodology to the human inducible NOS (iNOS) isoform, comparing wild-type (wt) and E546N mutant oxygenase/FMN (oxyFMN) constructs under near-native conditions. Using parallel reaction monitoring-based qXL MS, we assessed mu

SUBMITTER: Jiang T 

PROVIDER: S-EPMC12597282 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature