How mutations impression protein stability and construction dynamics is essential for understanding the molecular mechanism of the illness and the focused drug design. Nonetheless, probing the molecular particulars of mutation-induced delicate construction dynamics remains to be difficult.
A analysis group led by Prof. Wang Fangjun from the Dalian Institute of Chemical Physics (DICP) of the Chinese language Academy of Sciences has developed a time-resolved native mass spectrometry (TR-nMS) technique coupled with ultraviolet photodissociation (UVPD) evaluation. This technique can interrogate the mutation-induced delicate alterations in protein stability and construction unfolding dynamics. The research is printed within the Journal of the American Chemical Society.
The researchers initiated the protein unfolding course of by mixing the protein on-line with formic acid. With native mass spectrometry (nMS) and non-denaturing electrospray ionization (nESI), they monitored the species and relative intensities of acid-initiated protein unfolding intermediates through the distinctive cost state distributions (CSDs), and so they quantitatively characterised the M42T/H114R mutations induced stability alterations of goal proteins.
As well as, the researchers employed UVPD and fragment ion mass spectrometry strategies to quantitatively evaluate the dynamic construction and molecular particulars of the unfolding intermediates of wild-type dihydrofolate reductase (DHFR) and the mutant.
The UVPD evaluation revealed the particular stabilization impact of cofactor nicotinamide adenine dinucleotide phosphate (NADPH) on DHFR construction, and that the M42T/H114R mutations may scale back the non-covalent NADPH-DHFR interactions, together with residues I41, Q65, V78, D79, I82, and R98, thus selling a lower in stability.
“This work offers a brand new approach for learning the mutation-induced delicate construction dynamics and pathological mechanisms,” mentioned Prof. Wang.
Extra info:
Pan Luo et al, Time-Resolved Ultraviolet Photodissociation Mass Spectrometry Probes the Mutation-Induced Alterations in Protein Stability and Unfolding Dynamics, Journal of the American Chemical Society (2024). DOI: 10.1021/jacs.4c00316
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Chinese language Academy of Sciences
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New time-resolved ultraviolet photodissociation mass spectrometry technique for goal protein stability evaluation (2024, April 16)
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