Identification and partial characterisation of a chitinase from Nile tilapia, Oreochromis niloticus.
Measurement of chitinase activity in extracts from stomach, intestine, and serum of Nile tilapia with the artificial substrates 4-methylumbelliferil beta-D-N,N'-diacetylchitobioside and 4-methylumbelliferil beta-D-N,N'N"-triacetylchitotrioside (4MU[GlcNAc](2,3)) showed that an endochitinase was involved in the liberation of the fluorophore 4-methylumbelliferone (MU). Enzymes were isolated from tilapia serum by a combination of gel filtration, ion exchange, and reverse-phase chromatography. The molecular mass of the enzyme was estimated to be 75 kDa by SDS-PAGE, suggesting that the enzyme occurs as a monomer. The partially purified enzyme showed maximal activity at pH 7.0 when assayed with 4MU[GlcNAc](2) and lost its activity below pH 5.0 and above pH 8.0. The optimal pH of the purified enzyme toward the substrate 4MU[GlcNAc](3) was pH 9.0 and activity was lost below pH 8.0 and above pH 9.0. Our study has revealed the presence of a chitinolytic enzyme in the gastrointestinal tract and serum that may play a role in digestion and/or defense.
Molinari
Lígia Maria
LM
Programa de Pós-Graduação em Microbiologia, Universidade Estadual de Londrina, Brazil.
Pedroso
Raissa Bocchi
RB
Scoaris
Denise de Oliveira
Dde O
Ueda-Nakamura
Tânia
T
Nakamura
Celso Vataru
CV
Dias Filho
Benedito Prado
BP
D000818:Animals; D002688:Chitinases / Q000097:blood / Q000737:chemistry* / Q000302:isolation & purification; D023681:Cichlids / Q000097:blood*; D004789:Enzyme Activation / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D007422:Intestines / Q000201:enzymology; D013270:Stomach / Q000201:enzymology; D013379:Substrate Specificity / Q000502:physiology
D002688:Chitinases