Recovery and Characterization of a Serine Collagenolytic Extract from Snow Crab (Chionoecetes opilio) By-products

Abstract

Sequential acidic precipitation followed by a single chromatographic step (gel filtration) allowed the recovery of a collagenolytic fraction containing several proteases from by-products of snow crab (Chionoecetes opilio). The partial purification was particularly efficient to recover tryptic (purification fold = 1,352.5; yield = 110%) but also chymotryptic, elastolytic, and collagenolytic activities. A temperature of 40 °C and pH 8.0-8.5 were optimal for enzyme activity, which was stable for 2 h under these conditions. Calcium was not required for stability and thus activity. The isoelectric points of the protein components ranged from 3.7 to 4.6. Zymography revealed 29 and 48 kDa major components and others from 22 to 56 kDa. Enzymes were inhibited by PMSF and TLCK but were insensitive to TPCK. In view of these properties, the proteases likely belong to the serine collagenase group. Inhibition by EDTA could be due to a mechanism other than Ca(2+) chelation. Using a food system (ground fish), the fraction was more proteolytic than a commercial bacterial protease, suggesting potential applications in enzymatic hydrolysis processes.

Colagenase de pescada branca: extração, purificação parcial, caracterização e teste de especificidade ao colágeno para aplicação industrial

“…MUKHERJEE et al (2009) observed that pH 5.0 improved the activity of enzymes from species of sponge (R. odorabile). The stability of pH has also been reported for aquatic species such as fish, crustaceans, and sponges, and optimal values were found oscillating between < 5.0 and > 10.0 (TURKIEWIC et al, 1991;KRISTJÁMSSON et al, 1995;TERUEL and SIMPSON, 1995;MURADO et al, 2009;WU et al, 2010a;WU et al, 2010b;HAYET et al, 2011;SOUCHET and LAPLANTE, 2011;SUPHATHARAPRATEEP et al, 2011;BAEHAKI et al, 2012).…”

“…TERUEL andSIMPSON (1995), BYUN et al (2002), , SOUCHET andLAPLANTE (2011) andDABOOR et al (2012) reported an optimum pH of 7.5 to enzymes obtained from the aquatic species: winter flounder (P. americanus), tuna (T. thymus), Mackerel (S. japonicus), snow crab (Chionoecetes opilio) and fish waste, respectively. KRISTJÁMSSON et al (1995), ROY et al (1996) and BAEHAKI et al (2012) found an optimum pH of 7.0 for enzymes from Atlantic cod (G. morhua), greenshore crab (C. maenas) and Bacillus licheniformis, respectively.…”

“…Collagenases (EC 3. 4.24.7) constitute a group of enzymes that hydrolyze the peptide bonds of various types of collagen, the most abundant protein in mammals, under physiological conditions of pH and temperature both in vivo and in vitro (DABOOR et al, 2010;HAYET et al, 2011;SOUCHET and LAPLANTE, 2011). They are highly specific for native or denatured collagen, showing no activity to any other protein.…”

Study on enzymes of industrial interest in digestive viscera: Greater amberjack (Seriola dumerili)

“…Souchet and Laplante [28] also detected the collagenolytic activity (13.3 IU/mg) in byproducts of snow crab Chionoecetes opilio. Currently, much of the collagenase used in the market is of microbial origin.…”

Colagenasas do pescado y sus aplicaciones industriales

“…Para a maioria das espécies aquáticas, esse pH situa-se numa faixa entre 6,0 e 8,0 , Daboor et al, 2010. A estabilidade enzimática às variações de pH foi reportada para espécies aquáticas (peixes, crustáceos, esponjas), variando entre 5,0 a 10,0, a escala de recuperação de atividade (Kristjánsson et al, 1995, Teruel and Simpson, 1995, Lima et al, 2009, Mukherjee et al, 2009, Murado et al, 2009, Wu et al, 2010a, Hayet et al, 2011, Souchet and Laplante, 2011.…”