期刊
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
卷 1866, 期 11, 页码 -出版社
ELSEVIER
DOI: 10.1016/j.bbagen.2022.130217
关键词
Cyanophycin; Cyanophycinase; Diaminopropanoic acid; Acyl-enzyme intermediate; Structure Peptidase
资金
- National Institute of General Medical Sciences from the National Institutes of Health [P30 GM124165]
- DOE Office of Science by Argonne National Laboratory [DE-AC02-06CH11357]
- Canada Foundation for Innovation (CFI)
- Natural Sciences and Engineering Research Council (NSERC)
- National Research Council (NRC)
- Canadian Institutes of Health Research (CIHR)
- Government of Saskatchewan
- University of Saskatchewan
This study investigates the mechanism of how cyanophycinase specifically binds and degrades cyanophycin, a resistant polymer. The findings provide important insights into the metabolism of cyanophycin.
Background: Cyanophycinases are serine protease family enzymes which are required for the metabolism of cyanophycin, the natural polymer multi-L-arginyl-poly(L-aspartic acid). Cyanophycinases degrade cyanophycin to beta-Asp-Arg dipeptides, which enables use of this important store of fixed nitrogen. Methods: We used genetic code expansion to incorporate diaminopropionic acid into cyanophycinase in place of the active site serine, and determined a high-resolution structure of the covalent acyl-enzyme intermediate resulting from attack of cyanophycinase on a short cyanophycin segment. Results: The structure indicates that cyanophycin dipeptide residues P1 and P1' bind shallow pockets adjacent to the catalytic residues. We observe many cyanophycinase - P1 dipeptide interactions in the co-complex structure. Calorimetry measurements show that at least two cyanophycin dipeptides are needed for high affinity binding to cyanophycinase. We also characterized a putative cyanophycinase which we found to be structurally very similar but that shows no activity and could not be activated by mutation of its active site. General significance: Despite its peptidic structure, cyanophycin is resistant to degradation by peptidases and other proteases. Our results help show how cyanophycinase can specifically bind and degrade this important polymer.
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