The crystallographic structure of the subtilisin protease from Penicillium cyclopium.
The major extracellular protease from the fungus Pencillium cyclopium was crystallized in the presence of p-phenylmethanesulfonyl fluoride (PMSF) and investigated by X-ray diffraction analysis. It was subsequently cloned and the amino acid sequence deduced from its cDNA. Although the sequence is only 49% identical to that of proteinase K of Tritirachium album, the three-dimensional structures of the two proteases are virtually identical. The model for P. cyclopium protease was refined by simulated annealing to an R of 18% at 1.7 A resolution. The greatest variation from the proteinase K polypeptide is in loop 114-134 and is due to the absence of a disulfide bridge in the P. cyclopium protease that is present in proteinase K. A difference was also observed in the orientation of the histidine in the catalytic triad, though this could be due to the presence of PMSF at the active site. The coordination geometry of the strongly bound calcium in the P. cyclopium protease is octahedral and uses some different protein ligands than does proteinase K. In the protease from P. cyclopium there is no cysteine near the active site, nor is there a second calcium binding site as is found in proteinase K, suggesting that neither is important to catalytic activity.
Koszelak
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Department of Biochemistry, University of California, Riverside 92521, USA.
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Day
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Greenwood
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McPherson
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D000595:Amino Acid Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D018076:DNA, Complementary / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D019286:Endopeptidase K / Q000737:chemistry; D005658:Fungi / Q000201:enzymology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010407:Penicillium / Q000201:enzymology*; D010664:Phenylmethylsulfonyl Fluoride; D017433:Protein Structure, Secondary; D017385:Sequence Homology; D013381:Subtilisins / Q000737:chemistry*
D018076:DNA, Complementary; D004220:Disulfides; D010664:Phenylmethylsulfonyl Fluoride; D013381:Subtilisins; D019286:Endopeptidase K; D002118:Calcium