title: Digestion of the lambda cI repressor with various serine proteases and correlation with its three dimensional structure.
abstract: Partial proteolysis of the lambda cI repressor has been carried out systematically with trypsin, chymotrypsin, elastase, endoproteinase Glu-C, kallikrein, and thrombin. The cleavage sites have been determined by (i) comparison of fragments produced and observed in SDS-polyacrylamide gel with known fragments and plots of distance migrated versus log (molecular weight of fragment), (ii) partial Edman sequencing of the stable C-terminal fragments to identify cleavage points, and (iii) electrospray mass spectrometry of fragments produced. Most cleavage points are found to occur in the region 86-137, saving some in the N-terminal domain observed for trypsin and Glu-C. Region 86-137 can be further subdivided into three regions 86-91, 114-121, and 128-137 prone to cleavage, with intermediate regions resistant to cleavage to all six proteases. These resistant regions show that much of the region 93-131 previously called a 'linker' is actually part of the C-domain as first proposed in all models from our laboratory. Region 92-114 includes the cleavage site Ala-Gly, which must be buried in the intact repressor. The observed cleavage points in region 114-137 can be used to judge the best among three previously proposed models since they differ from each other in the structure of region 93-131. Model 1j5g is adjudged to be better than model 1lwq (which is based on 1kca, a crystal structure) as susceptible residues are more exposed in the former and lack of cleavages at six sites is better explained. Likewise, the models 1j5g and 1lwq are compared with a recent crystal structure of fragment 101-229 in 2ho0 and another low resolution crystal structure in 3bdn.
authors.affiliation: Department of Biochemistry, Bose Institute, P-1/12, C.I.T. Scheme VII M, Calcutta 700054, India.
authors.forename: Atasi
authors.initials: A
authors.lastname: Pal
authors.forename: Rajagopal
authors.initials: R
authors.lastname: Chattopadhyaya
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002918:Chymotrypsin / Q000378:metabolism; D007610:Kallikreins / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010196:Pancreatic Elastase / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D011487:Protein Conformation; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D012097:Repressor Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D007610:Kallikreins; D012697:Serine Endopeptidases; D002918:Chymotrypsin; D010196:Pancreatic Elastase; D013917:Thrombin