Cystatin like thiol proteinase inhibitor from pancreas of Capra hircus: purification and detailed biochemical characterization.
A thiol proteinase inhibitor from Capra hircus (goat) pancreas (PTPI) isolated by ammonium sulphate precipitation (20-80%) and gel filtration chromatography on Sephacryl S-100HR, with 20.4% yield and 500-fold purification, gave molecular mass of 44 kDa determined by its electrophoretic and gel filtration behavior, respectively. The stokes radius, diffusion and sedimentation coefficients of PTPI were 27.3 A, 7.87 x 10(-7) cm(2) s(-1) and 3.83 s, respectively. It was stable in pH range 3-10 and up to 70 degrees C (critical temperature, E (a) = 21 kJ mol(-1)). Kinetic analysis revealed reversible and competitive mode of inhibition with PTPI showing the highest inhibitory efficiency against papain (K ( i ) = 5.88 nM). The partial amino acid sequence analysis showed that it shared good homology with bovine parotid and skin cystatin C. PTPI possessed 17.18% alpha helical content assessed by CD spectroscopy. The hydropathy plot of first 24 residues suggested that most amino acids of this stretch might be in the hydrophobic core of the protein.
Priyadarshini
Medha
M
Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh, 202002, Uttar Pradesh, India.
Bano
Bilqees
B
D000595:Amino Acid Sequence; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D015891:Cystatins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D006041:Goats / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009928:Organ Specificity; D010179:Pancreas / Q000378:metabolism*; D055550:Protein Stability; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013438:Sulfhydryl Compounds / Q000032:analysis
D002241:Carbohydrates; D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D021122:Protein Subunits; D013438:Sulfhydryl Compounds; D003546:Cysteine Endopeptidases