Isolation and characterization of domain I of ovoinhibitor.
Domain I of ovoinhibitor was isolated by subjecting the protein to specific chemical cleavage by cyanogen bromide followed by repeated gel filtration. The first domain of ovoinhibitor was found to be homogeneous by the criteria of gel chromatography, SDS-PAGE and PAGE. Mr values by gel filtration (10900) and SDS-PAGE (8300) were slightly higher than that computed from amino-acid sequence. This discrepancy has been attributed to the glycoprotein nature of domain I as it was found to contain 10% neutral carbohydrate and 2% sialic acid. Fluorescence spectral properties showed the presence of tryptophan in domain I. The amino-acid composition of domain I isolated in this study was in very good agreement with that computed from amino-acid sequence. Gel filtration behaviour of the first domain was consistent with a Stokes radius of 1.6 nm and a frictional ratio of 1.2 suggesting asymmetry and/or excessive hydration. Domain I was found to be a potent inhibitor of bovine trypsin but was virtually devoid of activity against chymotrypsin, elastase and proteinase K. The equilibrium association constant for domain I-trypsin complex was computed to be 6.6x10(8)M-1.
Galzie
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Interdisciplinary Biotechnology Unit, J.N. Medical College, Aligarh Muslim University, Aligarh, India.
Qasim
M A
MA
Salahuddin
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A
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001665:Binding Sites; D002645:Chickens; D002850:Chromatography, Gel; D003488:Cyanogen Bromide; D015387:Egg Proteins, Dietary / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010059:Ovomucin / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D017186:Titrimetry; D014361:Trypsin Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology
D000596:Amino Acids; D015387:Egg Proteins, Dietary; D010446:Peptide Fragments; D011480:Protease Inhibitors; D014361:Trypsin Inhibitors; D010059:Ovomucin; C004224:ovoinhibitor; D003488:Cyanogen Bromide