A method for direct application of human plasmin on a dithiothreitol-containing agarose stacking gel system.
A new simplified procedure for identifying human plasmin was developed using a DTT copolymerized agarose stacking gel (ASG) system. Agarose (1 %) was used for the stacking gel because DTT inhibits the polymerization of acrylamide. Human plasmin showed the lowest activity at pH 9.0. There was a similar catalytically active pattern observed under acidic conditions (pH 3.0) to that observed under alkaline conditions (pH 10.0 or 11.0). Using the ASG system, the primary structure of the heavy chain could be established at pH 3.0. This protein was found to consist of three fragments, 45 kDa, 23 kDa, and 13 kDa. These results showed that the heavy chain has a similar structure to the autolysed plasmin (Wu et al., 1987b) but there is a different start amino acid sequence of the N-termini.
Choi
Nack-Shick
NS
Systemic Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejon 305-333.
Chung
Dong-Min
DM
Yoon
Kab-Seog
KS
Maeng
Pil Jae
PJ
Kim
Seung-Ho
SH
D000595:Amino Acid Sequence; D001671:Biochemistry / Q000379:methods*; D004220:Disulfides / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry / Q000494:pharmacology*; D004587:Electrophoresis, Agar Gel / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D005340:Fibrinogen / Q000737:chemistry; D005341:Fibrinolysin / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D010958:Plasminogen / Q000737:chemistry; D017434:Protein Structure, Tertiary; D012685:Sepharose / Q000737:chemistry; D013917:Thrombin / Q000737:chemistry
D004220:Disulfides; D005340:Fibrinogen; D010958:Plasminogen; D012685:Sepharose; D013917:Thrombin; D005341:Fibrinolysin; D004229:Dithiothreitol