Purification and characterization of a 34-kDa, heat stable glycoprotein from Synadenium grantii latex: action on human fibrinogen and fibrin clot.
Latex glycoprotein (LGP) from Synadenium grantii latex was purified by the combination of heat precipitation and gel permeation chromatography. LGP is a heat stable protein even at 80 degrees C showed a sharp single band both in SDS-PAGE as well as in native (acidic) PAGE. LGP is a monomeric protein appears as single band under reducing condition. It is a less hydrophobic protein showed sharp single peak in RP-HPLC with retention time of 13.3 m. The relative molecular mass of LGP is 34.4 kDa. CD spectrum of LGP explains less content of alpha-helix (7%), and high content of beta-pleated sheets (48%) and random coils (46%). The N-terminal sequence of LGP is D-F-P-S-D-W-Y-A-Y-E-G-Y-V-I-D-R-P-F-S. Purified LGP is a fibrinogen degrading protease hydrolyses all the three subunits in the order of Aalpha, Bbeta and gamma. The hydrolytic pattern is totally different from plasmin as well as thrombin. LGP reduces recalcification time from 165 to 30 s with citrated human plasma but did not show thrombin like as well as factor Xa-like activity. Although LGP induces procoagulant activity, it hydrolyses partially cross-linked fibrin clot. It hydrolyses all the subunits of partially cross-linked fibrin clot (alpha- chains, beta-chain and gamma-gamma dimer). LGP is a serine protease, inhibited by PMSF. Other serine protease inhibitors, aprotinin and leupeptin did not inhibit the caseinolytic activity as well as fibrinogenolytic activity. We report purification and characterization of a glycoprotein from Synadenium grantii latex with human fibrino(geno)lytic activity.
Rajesh
R
R
Department of Studies in Biochemistry, University of Mysore, Manasagangothri, Mysore 570006, India.
Nataraju
A
A
Gowda
C D R
CD
Frey
B M
BM
Frey
F J
FJ
Vishwanath
B S
BS
D000595:Amino Acid Sequence; D001777:Blood Coagulation / Q000187:drug effects*; D002942:Circular Dichroism; D019665:Euphorbiaceae / Q000145:classification / Q000201:enzymology*; D005337:Fibrin / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism*; D005342:Fibrinolysis; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D006801:Humans; D006868:Hydrolysis; D007840:Latex / Q000737:chemistry* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000378:metabolism; D012697:Serine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*
D006023:Glycoproteins; D007840:Latex; D011480:Protease Inhibitors; D005337:Fibrin; D005340:Fibrinogen; D012697:Serine Endopeptidases