Expression, purification and preliminary crystallographic analysis of dipeptidyl peptidase IV from Porphyromonas gingivalis.
The asaccharolytic periodontopathogen Porphyromonas gingivalis produces membrane-anchored proteases such as dipeptidyl peptidase IV that are involved in the destruction of host periodontal tissue. The extracellular domain of this enzyme was overexpressed in Escherichia coli as an N-terminal His-tag fusion protein, purified using standard metal-affinity chromatography and crystallized using the hanging-drop vapour-diffusion technique in 40% 2-methyl-2,4-pentanediol and 100 mM Tris-HCl pH 8.0. Diffraction data to 2.7 A resolution were collected using synchrotron radiation. The crystals belong to space group P2(1), with unit-cell parameters a = 117.0, b = 112.9, c = 310.0 A, beta = 95.0 degrees. There are ten molecules per asymmetric unit, indicating a solvent content of 50%. Data were also collected from selenomethionine-derived crystals and structure solution by SAD or MAD is in progress.
Rea
Dean
D
Department of Biological Sciences, University of Warwick, Gibbet Hill Road, Coventry CV4 7AL, England.
Lambeir
Anne Marie
AM
Kumagai
Yumi
Y
De Meester
Ingrid
I
Scharpé
Simon
S
Fülöp
Vilmos
V
D002845:Chromatography; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D016966:Porphyromonas gingivalis / Q000201:enzymology*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012645:Selenomethionine / Q000737:chemistry; D017356:Synchrotrons; D014961:X-Ray Diffraction
D011993:Recombinant Fusion Proteins; D012645:Selenomethionine; D018819:Dipeptidyl Peptidase 4