Identification of digestive hydrolases in the larval midgut of the camel nasal bot fly cephalopina titilla tor clark [diptera: Oestridae]

Activities of digestive hydrolases associated with midgut of the third instar larva of Cephalopina titillator were investigated. Based on the hydrolysis of synthetic substrates and optimum pH, it was found that C. titillator midgut contains trypsin-like [optimum pH, 9], chymotrypsin esterase-like [optimum pH, 8], carboxypeptidase A and B [optimum pH at 8.5 and 7 respectively], alkaline- and acidphosphatase [optimum pH at 9 and 5 respectively] and membrane bound leucine aminopeptidase [optimum pH, 8]. An acid proteinase activity was detected, by the ability to hydrolyze acid denaturated haemoglobin; and it seems to be close to pepsin than cathepsin-like enzyme. It has a maximum activity at pH 3.5. alpha-Glucosidase activity, and was also identified [optimum pH at 6] in the midgut, and seems to be membrane bound

Purification and characterization of a low-molecular-weight bovine kidney acid phosphatase

A relative low molecular mass bovine kidney acid phosphatase was purified 1,640-fold to homogeneity, with 7 percent recovery. The purified enzyme (specific activity 100 mumol min-1 mg-1) was electrophoretically homogeneous with a relative molecular massa of 17.8 kDa, as determined by SDS-polyacrylamide gel electrophoresis. A broad pH optimum of 4.0-5.5 and a maximal enzyme activity at 60 degrees Celsius were determined for the p-nitrophenyl phosphate hydrolysis. Apparent Km values of 0.14 mM, 0.4 mM, 0.3 mM and 7.9 mM were obtained, at 37 degrees Celsius and pH 5.0, for the best substrates p-nitrophenyl phosphate, beta-naphtyl-phosphate, flavin mononucleotide and tyrosine-phosphate, respectively. The enzyme activity was enhanced by guanosine but inhibited by ZnCl2 and CuSO4, p-cloromercuribenzoate and ammonium molybdate. Vanadate (Ki 0.47 muM), pyridoxal 5'-phosphate (Ki 2.2 muM), inorganic phosphate (Ki 0.77 mM) are competitive inhibitors. Both glycerol and methanol increased significantly the acide phosphatase activity, acting as good phosphate acceptors in the transphosphorylation reaction.

Unique structural features of red kidney bean purple acid phosphatase.

Purple acid phosphatase from red kidney beans (Phaseolus vulgaris) has been purified to homogeneity and characterized. The enzyme is a homodimer of 60 kDa subunits each containing one atom of zinc and iron in the active site. Circular dichroism spectral studies on the purified enzyme reveals that a large portion of the peptide backbone is in the unordered and beta-turn conformation. A unique feature of the red kidney bean acid phosphatase, which we have found, is that one of the two cysteines of each subunit is involved in the formation of an inter-subunit disulphide. The thiol group of the other cysteine is not necessary for the activity of the enzyme. Western blot analysis with antibodies raised against kidney bean acid phosphatase could not recognize acid phosphatases from other sources except from potato. This paper emphasizes the fact that acid phosphatases are functionally, but not structurally, conserved enzymes.