Crystallization of a soluble, catalytically active form of Escherichia coli leader peptidase.
Leader peptidase, a novel serine protease in Escherichia coli, catalyzes the cleavage of the amino-terminal leader sequences from exported proteins. It is an integral membrane protein containing two transmembrane segments with its carboxy-terminal catalytic domain residing in the periplasmic space. Here, we report a procedure for the purification and the crystallization of a soluble non-membrane-bound form of leader peptidase (delta 2-75). Crystals were obtained by the sitting-drop vapor diffusion technique using ammonium dihydrogen phosphate as the precipitant. Interestingly, we have found that the presence of the detergent Triton X-100 is required to obtain crystals sufficiently large for X-ray analysis. The crystals belong to the tetragonal space group P4(2)2(1)2, with unit cell dimensions of a = b = 115 A and c = 100 A, and contain 2 molecules per asymmetric unit. This is the first report of the crystallization of a leader (or signal) peptidase.
Paetzel
M
M
Department of Chemistry, Ohio State University, Columbus 43210, USA.
Chernaia
M
M
Strynadka
N
N
Tschantz
W
W
Cao
G
G
Dalbey
R E
RE
James
M N
MN
D003460:Crystallization; D018360:Crystallography, X-Ray; D010450:Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D008565:Membrane Proteins; D012697:Serine Endopeptidases; D012995:Solubility
D008565:Membrane Proteins; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase