Snake venom, diesterase enzymes

If a-casein is treated with either the crystalline pyrophosphatase of yeast at pH 7.0 (35) or with the snake venom diesterase at pH 8.2 (87), no inorganic phosphorus is released. However, if the diesterase reaction is carried out in weakly buffered solutions a small drop of pH takes place (71), indicating the exposure of acidic groups. Subsequent incubation of the diesterase-treated a-casein with prostate phosphatase at pH 6.0 hberates no more phosphorus than in the absence of the diesterase. If, how ever, prostate and intestinal phosphatase are added, 78 % of the a-casein phosphorus is set free. Since the intestinal enzyme at pH 6.0 acts on low... 

As is shown in Table VI, most of the phosphorus of /3-casein is liberated by the prostate enzyme at pH 6.0 after pretreatment with the snake venom diesterase. This indicates that the /3-protein contains diester bonds of the —0—P—0— type, which is in contrast to the —N—P—0— of the a-casein (71). 

The first RNA molecule to be sequenced was not a virus, but a tRNA. Two enzymes were required in these analyses (i) bovine pancreatic ribonuclease, which cleaved after pyrimidines (C or U), became a classic system for scientific studies after Armour Co., the hotdog Company, purified 1 kg of the enzyme and distributed it to scientists (source Wikipedia) and (ii) takadiastase ribonuclease Tl, which cleaved 3 to a guanosine (G). Each of these small fragments was further analyzed by partial digestion with snake venom diesterase from the 3 -ends. Once again, biochemistry and enzymology led to breakthroughs in chemical analysis. 

Desoxyribonucleic acid is hydrolyzed by calf intestinal diesterase to give a mixture of mononucleotides.Phosphatase activity present in the enzyme preparation must be inhibited by arsenate otherwise, nucleosides are obtained. The same nucleotide mixture results from the action of snake venom diesterase freed of phosphatase activity by chromatography on a cellulose column." ... 

Intestinal mucosa enzyme partially inhibited by arsenate leads to the formation of guanosine, cytidine, and cytidine-5 -phosphate. This preparation splits at (2) with subsequent partial dephosphorylation. Snake venom diesterase also sphts at (2). Prostatic phosphatase liberates... 

In addition to the proposed regulatory role of ATP and pyrophosphate, some possibility exists that 3, 5 -cyclic phosphate diesterase is under physiological control. Such ideas arose through observations of Cheung (43, 62) that the partially purified enzyme from beef brain was markedly activated by snake venom. The stimulatory factor was labile at extreme pH it was not dialyzable and appeared to be a protein. A similar activating factor is also present in brain tissue (63) and is removed during purification of the diesterase. It seems to interact stoichiometrically with the enzyme. The activator is destroyed by trypsin and is not proteolytic itself. The precise role of this protein in regulating the phosphodiesterase in vivo is not yet established, however. 

Venom Phosphodiesterase. A phosphodiesterase from the venom of several species of snakes exhibits extreme specificity with regard to nucleotides. It hydrolyzes only components from a phosphate esterified at a 5 position. Thus, 5 nucleotides are liberated from RNA. The venom diesterase attacks both purine and pyrimidine nucleotides (Fig. 26). This activity was useful in establishing that 3 -5 linkages, rather than 2 -3 linkages, occur in nucleic acids. A similar enzyme occurs in intestines. 

---