Purification, crystallization and preliminary X-ray diffraction studies to near-atomic resolution of dihydrodipicolinate synthase from methicillin-resistant Staphylococcus aureus.
In recent years, dihydrodipicolinate synthase (DHDPS; EC 4.2.1.52) has received considerable attention from both mechanistic and structural viewpoints. DHDPS is part of the diaminopimelate pathway leading to lysine, coupling (S)-aspartate-beta-semialdehyde with pyruvate via a Schiff base to a conserved active-site lysine. In this paper, the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of DHDPS from methicillin-resistant Staphylococcus aureus, an important bacterial pathogen, are reported. The enzyme was crystallized in a number of forms, predominantly from PEG precipitants, with the best crystal diffracting to beyond 1.45 A resolution. The space group was P1 and the unit-cell parameters were a = 65.4, b = 67.6, c = 78.0 A, alpha = 90.1, beta = 68.9, gamma = 72.3 degrees . The crystal volume per protein weight (V(M)) was 2.34 A(3) Da(-1), with an estimated solvent content of 47% for four monomers per asymmetric unit. The structure of the enzyme will help to guide the design of novel therapeutics against the methicillin-resistant S. aureus pathogen.
Burgess
Benjamin R
BR
Department of Biochemistry and Molecular Biology, University of Melbourne, Parkville, Victoria 3010, Australia.
Dobson
Renwick C J
RC
Dogovski
Con
C
Jameson
Geoffrey B
GB
Parker
Michael W
MW
Perugini
Matthew A
MA
D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D003460:Crystallization; D006836:Hydro-Lyases / Q000737:chemistry* / Q000302:isolation & purification; D016106:Methicillin Resistance; D018625:Microscopy, Atomic Force; D011092:Polyethylene Glycols; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology* / Q000472:pathogenicity; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D011092:Polyethylene Glycols; D006836:Hydro-Lyases; C020040:4-hydroxy-tetrahydrodipicolinate synthase