Korean J Parasitol.  1999 Mar;37(1):39-46. 10.3347/kjp.1999.37.1.39.

A 54 kDa cysteine protease purified from the crude extract of Neodiplostomum seoulense adult worms

• Choi MH
• Choe SC
• Lee SH

• KMID: 1715282
• DOI: http://doi.org/10.3347/kjp.1999.37.1.39

Abstract

As a preliminary study for the explanation of pathobiology of Neodiplostomum seoulense infection, a 54 kDa protease was purified from the crude extract of adult worms by sequential chromatographic methods. The crude extract was subjected to DEAE-Sepharose Fast Flow column, and protein was eluted using 25 mM Tris-HCl (pH 7.4) containing 0.05, 0.1, 0.2 and 0.4 M NaCl in stepwise elution. The 0.2 M NaCl fraction was further purified by Q-Sepharose chromatography and protein was eluted using 20 mM sodium acetate (pH 6.4) containing 0.05, 0.1, 0.2 and 0.3 M NaCl, respectively. The 0.1M NaCl fraction showed a single protein band on SDS-PAGE carried out on a 7.5-15% gradient gel. The proteolytic activities of the purified enzyme were specifically inhibited by L-trans-epoxy-succinylleucylamide (4-guanidino) butane (E-64) and iodoacetic acid. The enzyme, cysteine protease, showed the maximum proteolytic activity at pH 6.0 in 0.1 M buffer, and degraded extracellular matrix proteins such as collagen and fibronectin with different activities. It is suggested that the cysteine protease may play a role in the nutrient uptake of N. seoulense from the host intestine.

MeSH Terms

Animal
Cattle
Cysteine Endopeptidases/physiology
Cysteine Endopeptidases/isolation & purification*
Extracellular Matrix Proteins/metabolism
Host-Parasite Relations
Human
Molecular Weight
Rats
Rats, Sprague-Dawley
Snakes
Trematoda/pathogenicity
Trematoda/enzymology*