scholarly journals Purification and characterization of a high-Mr proteinase inhibitor of pro-phenol oxidase activation from crayfish plasma

1987 ◽  
Vol 248 (1) ◽  
pp. 223-228 ◽  
Author(s):  
H G Hergenhahn ◽  
A Aspan ◽  
K Söderhäll
Keyword(s):  
Proteinase Inhibitor ◽  
Serine Proteinase ◽  
Hydrophobic Interaction Chromatography ◽  
Phenol Oxidase ◽  
Purification And Characterization ◽  
Recognition Mechanism ◽  
Heat Stable ◽  
Apparent Homogeneity ◽  
Activation Cascade

Crayfish plasma was found to contain a proteinase inhibitor, which was purified to apparent homogeneity by acid precipitation, affinity chromatography on concanavalin A-Sepharose and hydrophobic-interaction chromatography. The inhibitor is a monomeric protein with an Mr of about 155,000 and a pI in the range 4.6-4.8. It is heat-stable and tolerant to low pH. It inhibits the serine proteinases trypsin and chymotrypsin, but not thrombin or subtilisin. Furthermore, it is efficient in decreasing the activity of a proteinase from crayfish haemolymph that is involved in the activation cascade of pro-phenol oxidase and can also block pro-phenol oxidase activation by this serine proteinase. This cascade is believed to play a central role in the recognition mechanism of non-self material in crustaceans and insects. The data presented give some evidence that the new proteinase inhibitor is involved in the regulation of this system.

scholarly journals Purification and characterization of an α2-macroglobulin-like proteinase inhibitor from plasma of the crayfish Pacifastacus leniusculus

1988 ◽  
Vol 255 (3) ◽  
pp. 801-806 ◽  
Author(s):  
H G Hergenhahn ◽  
M Hall ◽  
K Söderhäll
Keyword(s):  
Proteinase Inhibitor ◽  
Hydrophobic Interaction Chromatography ◽  
Acid Precipitation ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Pacifastacus Leniusculus ◽  
Purification And Characterization ◽  
Apparent Homogeneity ◽  
Α2 Macroglobulin

An alpha 2-macroglobulin (alpha 2M)-like proteinase inhibitor from plasma of the crayfish Pacifastacus leniusculus was purified to apparent homogeneity by acid precipitation, hydrophobic interaction chromatography, affinity chromatography on concanavalin A-Sepharose and anion-exchange chromatography. The subunit Mr is about 190,000. Pore-size-limit electrophoresis proved the native protein to be a dimer. The purified protein resembled vertebrate alpha 2 Ms in that it protected trypsin from inhibition by soyabean trypsin inhibitor, and in its sensitivity to methylamine treatment. Methylamine also prevented the protein from being autolytically cleaved into Mr 60,000 and 140,000 fragments when subjected to heat treatment. The amino acid composition showed similarities with both human alpha 2 M and an alpha 2 M-like protein from the arthropod Limulus polyphemus. These data indicate that this Pacifastacus alpha 2M-like protein (P alpha 2M) may be a distantly related homologue of vertebrate alpha 2Ms.

scholarly journals Proteinase inhibitors in rat serum. Purification and partial characterization of three functionally distinct trypsin inhibitors

1984 ◽  
Vol 218 (3) ◽  
pp. 953-959 ◽  
Author(s):  
L Kuehn ◽  
M Rutschmann ◽  
B Dahlmann ◽  
H Reinauer
Keyword(s):  
Proteinase Inhibitor ◽  
Proteinase Inhibitors ◽  
Serine Proteinase ◽  
The Other ◽  
Serine Proteinases ◽  
Rat Serum ◽  
Apparent Homogeneity ◽  
Human Counterpart ◽  
Proteinase Inhibitor Ii

Three different serine proteinase inhibitors were isolated from rat serum and purified to apparent homogeneity. One of the inhibitors appears to be homologous to alpha 1-proteinase inhibitor isolated from man and other species, but the other two, designated rat proteinase inhibitor I and rat proteinase inhibitor II, seem to have no human counterpart. alpha 1-Proteinase inhibitor (Mr 55000) inhibits trypsin, chymotrypsin and elastase, the three serine proteinases tested. Rat proteinase inhibitor I (Mr 66000) is active towards trypsin and chymotrypsin, but is inactive towards elastase. Rat proteinase inhibitor II (Mr 65000) is an effective inhibitor of trypsin only. Their contributions to the trypsin-inhibitory capacity of rat serum are about 68, 14 and 18% for alpha 1-proteinase inhibitor, rat proteinase inhibitor I and rat proteinase inhibitor II respectively.

scholarly journals Purification and characterization of the heat-stable serine proteinase from Thermomonospora fusca YX

1987 ◽  
Vol 246 (2) ◽  
pp. 511-517 ◽  
Author(s):  
T W Gusek ◽  
J E Kinsella
Keyword(s):  
Thermal Stability ◽  
Ionic Strength ◽  
Serine Proteinase ◽  
Maximum Activity ◽  
Exchange Chromatography ◽  
Cation Exchange Chromatography ◽  
Purification And Characterization ◽  
Thermomonospora Fusca ◽  
Heat Stable

The proteinase secreted from Thermomonospora fusca YX grown on cellulose was purified by (NH4)2SO4 fractionation and cation-exchange chromatography. The isolated proteinase readily hydrolysed several proteins and demonstrated activity towards casein from 35 to 95 degrees C (at pH 8.0) with maximum activity at 80 degrees C. It exhibited broad pH and ionic-strength optima centered at pH 9.0 and 0.2 M-NaCl respectively, and it retained high activity in the presence of 2% (w/v) SDS, 20 mM-dithiothreitol and 1.0 M-NaCl. The proteinase, which was fully inhibited by phenylmethanesulphonyl fluoride, had an Mr of 14,500 and an isoelectric point at 9.21. A measurement of proteinase thermal stability demonstrated a T50% (15 min) of 85 degrees C at pH 4.5.

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