3EN0

The Structure of Cyanophycinase

Summary for 3EN0

• Entry DOI: 10.2210/pdb3en0/pdb
• Related: 1fy2 1fye
• Descriptor: Cyanophycinase, SULFATE ION (3 entities in total)
• Functional Keywords: serine protease, beta peptide specific, hydrolase, protease
• Biological source: Synechocystis sp.
• Total number of polymer chains: 3
• Total formula weight: 95076.16

Authors

Kimber, M.S.,Law, A.M.,Lai, S.W.S.,Tavares, J. (deposition date: 2008-09-25, release date: 2009-07-28, Last modification date: 2024-02-21)

Primary citation

Law, A.M.,Lai, S.W.,Tavares, J.,Kimber, M.S.
The structural basis of beta-peptide-specific cleavage by the serine protease cyanophycinase.
J.Mol.Biol., 392:393-404, 2009

PubMed Abstract: Cyanophycin, or poly-L-Asp-multi-L-Arg, is a non-ribosomally synthesized peptidic polymer that is used for nitrogen storage by cyanobacteria and other select eubacteria. Upon synthesis, it self-associates to form insoluble granules, the degradation of which is uniquely catalyzed by a carboxy-terminal-specific protease, cyanophycinase. We have determined the structure of cyanophycinase from the freshwater cyanobacterium Synechocystis sp. PCC6803 at 1.5-A resolution, showing that the structure is dimeric, with individual protomers resembling aspartyl dipeptidase. Kinetic characterization of the enzyme demonstrates that the enzyme displays Michaelis-Menten kinetics with a k(cat) of 16.5 s(-1) and a k(cat)/K(M) of 7.5x10(-6) M(-1) s(-1). Site-directed mutagenesis experiments confirm that cyanophycinase is a serine protease and that Gln101, Asp172, Gln173, Arg178, Arg180 and Arg183, which form a conserved pocket adjacent to the catalytic Ser132, are functionally critical residues. Modeling indicates that cyanophycinase binds the beta-Asp-Arg dipeptide residue immediately N-terminal to the scissile bond in an extended conformation in this pocket, primarily recognizing this penultimate beta-Asp-Arg residue of the polymeric chain. Because binding and catalysis depend on substrate features unique to beta-linked aspartyl peptides, cyanophycinase is able to act within the cytosol without non-specific cleavage events disrupting essential cellular processes.

PubMed: 19591842
DOI: 10.1016/j.jmb.2009.07.001

Experimental method

X-RAY DIFFRACTION (1.5 Å)