Expression, purification, and characterization of diacylated Lipo-YcjN from Escherichia coli

Treviño, Matthew A.; Amankwah, Kofi A.; Fernandez, Daniel; Weston, Scott A.; Stewart, Claire J.; Gallardo, Jaime Morales; Shahgholi, Mona; Sharaf, Naima G.

doi:10.1016/j.jbc.2024.107853

Title: Expression, purification, and characterization of diacylated Lipo-YcjN from Escherichia coli

Journal Article · 30 September 2024 · Journal of Biological Chemistry

DOI: https://doi.org/10.1016/j.jbc.2024.107853 · OSTI ID:2452850

^[1]; Amankwah, Kofi A. ^[1];

^[1]; Weston, Scott A. ^[1]; Stewart, Claire J. ^[1]; Gallardo, Jaime Morales ^[1]; Shahgholi, Mona ^[2];

^[1]

Stanford Univ., CA (United States)
California Institute of Technology (CalTech), Pasadena, CA (United States)

YcjN is a putative substrate binding protein expressed from a cluster of genes involved in carbohydrate import and metabolism in Escherichia coli. Here, we determine the crystal structure of YcjN to a resolution of 1.95 Å, revealing that its three-dimensional structure is similar to substrate binding proteins in subcluster D-I, which includes the well-characterized maltose binding protein. Furthermore, we found that recombinant overexpression of YcjN results in the formation of a lipidated form of YcjN that is posttranslationally diacylated at cysteine 21. Comparisons of size-exclusion chromatography profiles and dynamic light scattering measurements of lipidated and nonlipidated YcjN proteins suggest that lipidated YcjN aggregates in solution via its lipid moiety. Additionally, bioinformatic analysis indicates that YcjN-like proteins may exist in both Bacteria and Archaea, potentially in both lipidated and nonlipidated forms. Together, our results provide a better understanding of the aggregation properties of recombinantly expressed bacterial lipoproteins in solution and establish a foundation for future studies that aim to elucidate the role of these proteins in bacterial physiology.

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Research Organization:: SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)

Sponsoring Organization:: National Institutes of Health (NIH); USDOE Office of Science (SC), Basic Energy Sciences (BES); USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: AC02-76SF00515

OSTI ID:: 2452850

Journal Information:: Journal of Biological Chemistry, Journal Name: Journal of Biological Chemistry Journal Issue: 11 Vol. 300; ISSN 0021-9258

Publisher:: American Society for Biochemistry and Molecular BiologyCopyright Statement

Country of Publication:: United States

Language:: English

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