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Dissecting diazirine photo-reaction mechanism for protein residue-specific cross-linking and distance mapping.
Jiang, Yida; Zhang, Xinghe; Nie, Honggang; Fan, Jianxiong; Di, Shuangshuang; Fu, Hui; Zhang, Xiu; Wang, Lijuan; Tang, Chun.
Afiliação
  • Jiang Y; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Zhang X; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Nie H; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Fan J; Analytical Instrumentation Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Di S; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Fu H; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Zhang X; Analytical Instrumentation Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Wang L; Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • Tang C; Analytical Instrumentation Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
Nat Commun ; 15(1): 6060, 2024 Jul 18.
Article em En | MEDLINE | ID: mdl-39025860
ABSTRACT
While photo-cross-linking (PXL) with alkyl diazirines can provide stringent distance restraints and offer insights into protein structures, unambiguous identification of cross-linked residues hinders data interpretation to the same level that has been achieved with chemical cross-linking (CXL). We address this challenge by developing an in-line system with systematic modulation of light intensity and irradiation time, which allows for a quantitative evaluation of diazirine photolysis and photo-reaction mechanism. Our results reveal a two-step pathway with mainly sequential generation of diazo and carbene intermediates. Diazo intermediate preferentially targets buried polar residues, many of which are inaccessible with known CXL probes for their limited reactivity. Moreover, we demonstrate that tuning light intensity and duration enhances selectivity towards polar residues by biasing diazo-mediated cross-linking reactions over carbene ones. This mechanistic dissection unlocks the full potential of PXL, paving the way for accurate distance mapping against protein structures and ultimately, unveiling protein dynamic behaviors.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Reagentes de Ligações Cruzadas / Diazometano Idioma: En Revista: Nat Commun Assunto da revista: BIOLOGIA / CIENCIA Ano de publicação: 2024 Tipo de documento: Article País de afiliação: China

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Reagentes de Ligações Cruzadas / Diazometano Idioma: En Revista: Nat Commun Assunto da revista: BIOLOGIA / CIENCIA Ano de publicação: 2024 Tipo de documento: Article País de afiliação: China