2ZEG

Crystal structure of the human glutaminyl cyclase mutant E201L at 2.08 angstrom resolution

Summary for 2ZEG

• Entry DOI: 10.2210/pdb2zeg/pdb
• Related: 2AFM 2ZED 2ZEE 2ZEF 2ZEH 2ZEL 2ZEM 2ZEN 2ZEO 2ZEP
• Descriptor: Glutaminyl-peptide cyclotransferase, ZINC ION, SULFATE ION, ... (5 entities in total)
• Functional Keywords: hydrogen bond network, glutaminyl cyclase, pyroglutamate, site-directed mutagenesis, proton transfer, acyltransferase, glycoprotein, metal-binding, transferase
• Biological source: Homo sapiens (human)
• Cellular location: Secreted: Q16769
• Total number of polymer chains: 2
• Total formula weight: 75544.00

Authors

Huang, K.F.,Wang, Y.R.,Chang, E.C.,Chou, T.L.,Wang, A.H. (deposition date: 2007-12-12, release date: 2008-04-22, Last modification date: 2023-11-01)

Primary citation

Huang, K.F.,Wang, Y.R.,Chang, E.C.,Chou, T.L.,Wang, A.H.
A conserved hydrogen-bond network in the catalytic centre of animal glutaminyl cyclases is critical for catalysis.
Biochem.J., 411:181-190, 2008

PubMed Abstract: QCs (glutaminyl cyclases; glutaminyl-peptide cyclotransferases, EC 2.3.2.5) catalyse N-terminal pyroglutamate formation in numerous bioactive peptides and proteins. The enzymes were reported to be involved in several pathological conditions such as amyloidotic disease, osteoporosis, rheumatoid arthritis and melanoma. The crystal structure of human QC revealed an unusual H-bond (hydrogen-bond) network in the active site, formed by several highly conserved residues (Ser(160), Glu(201), Asp(248), Asp(305) and His(319)), within which Glu(201) and Asp(248) were found to bind to substrate. In the present study we combined steady-state enzyme kinetic and X-ray structural analyses of 11 single-mutation human QCs to investigate the roles of the H-bond network in catalysis. Our results showed that disrupting one or both of the central H-bonds, i.e., Glu(201)...Asp(305) and Asp(248)...Asp(305), reduced the steady-state catalysis dramatically. The roles of these two COOH...COOH bonds on catalysis could be partly replaced by COOH...water bonds, but not by COOH...CONH(2) bonds, reminiscent of the low-barrier Asp...Asp H-bond in the active site of pepsin-like aspartic peptidases. Mutations on Asp(305), a residue located at the centre of the H-bond network, raised the K(m) value of the enzyme by 4.4-19-fold, but decreased the k(cat) value by 79-2842-fold, indicating that Asp(305) primarily plays a catalytic role. In addition, results from mutational studies on Ser(160) and His(319) suggest that these two residues might help to stabilize the conformations of Asp(248) and Asp(305) respectively. These data allow us to propose an essential proton transfer between Glu(201), Asp(305) and Asp(248) during the catalysis by animal QCs.

PubMed: 18072935
DOI: 10.1042/BJ20071073

Experimental method

X-RAY DIFFRACTION (2.08 Å)