Scientists develop new techniques for in situ resolution of protein complexes

As performers of life activities, proteins perform their specific biological functions, for example, by interacting to form complexes. Recently, Zhang Lihua, a researcher at the Dalian Institute of Chemical Physics, Chinese Academy of Sciences, and Zhao Qun, a researcher at the Dalian Institute of Chemical Physics, Chinese Academy of Sciences, developed a mass spectrometry fragmentable crosslinker based on glycosidic bonds, which significantly improved the retrieval throughput and identification accuracy of crosslinking information, and at the same time had good amphiphilia and biocompatibility, realizing in situ crosslinking and large-scale accurate analysis of protein complexes in living cells. The results were published in the German Journal of Applied Chemistry.

Photo courtesy of Dalian Chemical Properties

Intracellular confinement effects, crowding effects, and organelle microenvironments are critical for maintaining protein complex structure and function. Chemical crosslinking technology, especially in situ chemical crosslinking mass spectrometry, has the advantage of large-scale analysis of in situ conformation and interaction interface of protein complexes, and has become an important technology for the analysis of protein complexes in living cells. However, at present, in situ crosslinking of living cells faces problems such as large cell perturbation and high complexity of cross-linked peptide spectra. Therefore, how to achieve in situ rapid crosslinking under low perturbation of living cells is a prerequisite for accurate protein in situ conformation and interaction analysis.

In this work, based on the high biocompatibility of sugar molecules and the fragmentable characteristics of glycosidic bonds, the team introduced glycosidic bonds into the skeleton design of functional crosslinkers, screened and obtained trehalose with high biocompatibility as the skeleton molecule, and developed a mass spectrometry fragmentable crosslinker-trehalose disuccinimidyl ester. Compared with the currently reported permeable membrane chemical crosslinkers, this crosslinker demonstrates better cell viability to maintain cell viability and can achieve efficient crosslinking of intracellular protein complexes in a low perturbation state.

On this basis, the selective fragmentation mode of mass spectrometry of low-energy glycosidic bonds-high-energy peptide bonds can reduce the data analysis of “I-shaped” cross-linked peptides to linear peptide data retrieval modified by conventional crosslinker fragments, which greatly reduces the complexity of cross-linked peptide spectral analysis and significantly improves the identification efficiency and accuracy of crosslinked peptides. (Source: Sun Danning, China Science News)

Related paper information:

Source link

Related Articles

Leave a Reply

Your email address will not be published. Required fields are marked *

Back to top button