Time-Resolved Ultraviolet Photodissociation Mass Spectrometry Probes the Mutation-Induced Alterations in Protein Stability and Unfolding Dynamics.
J Am Chem Soc
; 146(13): 8832-8838, 2024 Apr 03.
Article
em En
| MEDLINE
| ID: mdl-38507251
ABSTRACT
How mutations impact protein stability and structure dynamics is crucial for understanding the pathological process and rational drug design. Herein, we establish a time-resolved native mass spectrometry (TR-nMS) platform via a rapid-mixing capillary apparatus for monitoring the acid-initiated protein unfolding process. The molecular details in protein structure unfolding are further profiled by a 193 nm ultraviolet photodissociation (UVPD) analysis of the structure-informative photofragments. Compared with the wild-type dihydrofolate reductase (WT-DHFR), the M42T/H114R mutant (MT-DHFR) exhibits a significant stability decrease in TR-nMS characterization. UVPD comparisons of the unfolding intermediates and original DHFR forms indicate the special stabilization effect of cofactor NADPH on DHFR structure, and the M42T/H114R mutations lead to a significant decrease in NADPH-DHFR interactions, thus promoting the structure unfolding. Our study paves the way for probing the mutation-induced subtle changes in the stability and structure dynamics of drug targets.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Escherichia coli
/
Desdobramento de Proteína
Idioma:
En
Revista:
J Am Chem Soc
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
China