The structure of cyanophycinase in complex with a cyanophycin degradation intermediate.

Bacterial Proteins Dipeptides Peptide Hydrolases Polymers
Acyl-enzyme intermediate Cyanophycin Cyanophycinase Diaminopropanoic acid Peptidase Structure

Journal

Biochimica et biophysica acta. General subjects
ISSN: 1872-8006
Titre abrégé: Biochim Biophys Acta Gen Subj
Pays: Netherlands
ID NLM: 101731726

Informations de publication

Date de publication:
11 2022
Historique:
received: 19 05 2022
revised: 07 07 2022
accepted: 22 07 2022
pubmed: 30 7 2022
medline: 31 8 2022
entrez: 29 7 2022
Statut: ppublish

Résumé

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 β-Asp-Arg dipeptides, which enables use of this important store of fixed nitrogen. 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. 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. 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.

Sections du résumé

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 β-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.

Identifiants

DOI: 10.1016/j.bbagen.2022.130217 PMID: 35905922
pubmed: 35905922
pii: S0304-4165(22)00135-0
doi: 10.1016/j.bbagen.2022.130217
pii:
doi:

Substances chimiques

Bacterial Proteins 0
Dipeptides 0
Polymers 0
cyanophycin 0
Peptide Hydrolases EC 3.4.-
cyanophycinase EC 3.4.-

Types de publication

Journal Article Research Support, Non-U.S. Gov't

Langues

eng

Sous-ensembles de citation

IM

Pagination

130217

Subventions

Organisme : CIHR
ID : 178084
Pays : Canada

Informations de copyright

Copyright © 2022 Elsevier B.V. All rights reserved.

Auteurs

Itai Sharon (I)

Department of Biochemistry and Centre de recherche en biologie structurale, McGill University, Montréal, QC H3G 0B1, Canada.

Marcel Grogg (M)

Laboratory of Organic Chemistry, ETH Zürich, CH-8093 Zürich, Switzerland.

Donald Hilvert (D)

Laboratory of Organic Chemistry, ETH Zürich, CH-8093 Zürich, Switzerland.

T Martin Schmeing (TM)

Department of Biochemistry and Centre de recherche en biologie structurale, McGill University, Montréal, QC H3G 0B1, Canada. Electronic address: martin.schmeing@mcgill.ca.

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Classifications MeSH

questionsmedicales.fr › Acides aminés, peptides et protéines › Protéines › Protéines bactériennes › The structure of cyanophycinase in complex...
questionsmedicales.fr › Acides aminés, peptides et protéines › Peptides › Oligopeptides › Dipeptides › The structure of cyanophycinase in complex...
questionsmedicales.fr › Enzymes et coenzymes › Enzymes › Hydrolases › Peptide hydrolases › The structure of cyanophycinase in complex...
questionsmedicales.fr › Technologie, industrie et agriculture › Produits manufacturés › Matériaux biomédicaux et dentaires › Polymères › The structure of cyanophycinase in complex...