Adaptation of trypsin, chymotrypsin and α-amylase to casein level and protein source in Penaeus vannamei (Crustacea Decapoda)

Abstract

Adaptation of digestive enzymes to dietary proteins was studied in Penaeus vannamei at the level of enzyme activity, synthesis measured by an immunoquantitative assay and polymorphism estimated for enzymes by electrophoresis and for mRNA by RT-PCR amplification. An initial experiment was conducted over 20 days with dietary casein increasing between 25 and 48%, starch and the other constituants remaining unchanged. A significant dose response effect was established between trypsin and casein ranging from 25 to 40% in the diet. No apparent change in the isoform pattern was recorded. Chymotrypsin varied neither at the level of specific activities nor at the level of polymorphism. Amylase activities and amounts decreased while casein increased. Two major isoforms were detected for 25% casein in the diet and only one for 40% casein. In order to establish the level of polymorphism of the expressed amylase mRNAs, a 378 bp fragment was obtained by RT-PCR on total RNA. In the case of the 25% casein diet, two different fragments, which corresponded to two proteins differing by an acidic charge, were amplified while only one was amplified in the case of the 40% casein diet. A comparison of casein with other sources of protein, gelatin, squid meal and fish protein soluble concentrate at two different concentrations in the diets (25 and 40%, respectively) confirmed the stimulation of trypsin by casein. No dose response was measured with other protein sources: moreover, an inhibition of trypsin activity was measured with gelatin in the diet. Highest activities of chymotrypsin were measured with casein and squid meal diets and lowest activities with gelatin and FPSC diets. Concerning amylase, the same variations were measured. Moreover, all the 40% protein diets inhibited the expression of one isoform.