Partial Protease Characterization, In Vitro Protein Digestibility And Utilization Of Dietary Soybean In Giant Freshwater Prawn, Macrobrachium Rosenbergii (De Man)
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Date
2005-05
Authors
Chisty, Md. Ayaz Hasan
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Journal ISSN
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Publisher
Universiti Sains Malaysia
Abstract
Biochemical assays and electrophoretical observations using SDS-PAGE was
conducted for partial identification and characterization of proteases in the digestive
system of giant freshwater prawn (Macrobrachium rosenbergii). pH optima of the crude
enzyme extracts of prawn showed 3 peaks in activity at pH 3.0, 6.0 and 9.0. The acid
proteinase and trypsin at pH 2.5 and 9.0 showed higher activity respectively at 3 and
5hrs after the last feeding. Presence of serine class proteinase was identified using
PMSF that reduced 33% of the total proteolytic activity. Reduction in activity by 82%
and 55% using specific inhibitors TLCK and TPCK confirmed the presence of trypsin
and chymotrypsin respectively while EDTA reduced 34% activity resembles the
presence of metallo-proteinase. Characterization of the alkaline proteinases by 12%
SDS-PAGE, after incubation of extract with or without specific inhibitors, produced six
active bands of 13 to 136 kDa molar mass. Two trypsin bands of 13 and 36 kDa, three
chymotrypsin bands of 23, 47 and 73 kDa and one metallo-proteinase of 136 kDa were identified from zymogram. The results suggest that an acid protease responsible for the
initial digestion of proteins and trypsin and chymotrypsin are the main enzymes responsible for the rest.
Two in vitro techniques, viz. Lazo single enzyme assay using bovine trypsin and prawn crude enzyme assay, were used to measure the relative protein digestibility
(RPD) of five feed ingredients namely fishmeal (FM), extracted soybean meal (ExSM),mustard oil cake (MOC), sesame oil cake (SOC) and coconut oil cake (COC) and
seven practical diets. The RPD of feed ingredients and diets in Lazo single enzyme
assay ranged between 44-60% and 42-53% and that of prawn crude enzyme were 34-
82% and 74-84%. However the results between these two methods were contradictory
in that the RPD values were found to be higher in prawn enzyme but lower when Lazo enzyme system was used.
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Keywords
Natural history