Metal ion acceleration of the conversion of trypsinogen to trypsin. Lanthanide ions as calcium ion substitutes

Abstract

Further, as shown in Table I, not all of the hydrogen peroxide in the steady-state system is utilized to oxidize ferrocyanide at pH 7.8. In addition it has been observed that a large excess of hydrogen peroxide added to cytochrome c peroxidase at pH 8 causes rapid destruction of the heme group.The above results suggest that at high pH, ferrocyanide is such a poor substrate for I and I1 that hydrogen peroxide can effectively compete with ferrocyanide in reactions with these enzyme intermediates, producing a number of side reactions including oxidation of the heme and amino acid residues of the protein. During the side reactions, an intermediate is most likely produced which is a better oxidant of ferrocyanide than 11. This could account for the faster oxidation of ferrocyanide in the steady state compared to the single turnover transient state studies where only I and I1 are produced. p H Dependence. The pH dependence of the steady-state parameters is difficult to interpret due to the specific ion effects, the abnormal ionic strength dependence of VmaPP/e at pH 6, and the side reactions between enzyme and hydrogen peroxide at high pH. Certainly the pH dependence of VmaPP/eKmaPP, after correction for the side reactions, is consistent with the interpretation of and k3aPP in the transient state (Jordi and Erman, 1974). The pH dependence is due primarily to the variation in electrostatic interaction between ferrocyanide and the enzyme intermediates as the net charge on the enzyme changes with pH.Vmapp/e, essentially a unimolecular rate constant, should be relatively insensitive to electrostatic effects although the ionic strength dependence of Vmapp/e at pH 6 casts some doubt on this expectation. Nevertheless, if V,app/e is insensitive to the charge on the enzyme, then the strong pH dependence of VmaPP/e suggests that a proton or a protonated form of the complex may be involved in the intramolecular electron transfer reaction. This is an interesting area for further investigation.Kmapp is relatively independent of pH, varying by only a factor of 10 over the pH range 4-8. This is due to the cancellation of the pH variation of the intramolecular electron-transfer rate constants in the numerator and the pH dependence of the bimolecular association rate constants in the denominator of the expression for KmaPP, eq 7.ABSTRACT: The lanthanide ions are shown to be effective calcium ion substitutes in accelerating the conversion of trypsinogen to trypsin. The rate of activation of the zymogen in the presence of lanthanides is much greater than that of the calcium ion. In addition this increased activation takes place at nearly 100-fold lower concentrations of the lanthanide ions than with calcium ion. The effect of the lanthanide ions in accelerating the activation of the zymogen, like the calcium efw t h i n the last I O years there has been substantial attention focused on specific interactions of proteins with metal ions. It has become apparent that the functions of metal ion-protein complexes are biologically important not...

Substrate recognition and catalysis by GH47 α-mannosidases involved in Asn-linked glycan maturation in the mammalian secretory pathway

“…A consequence of this attack would be the transient change from an eight to seven coordinated Ca 2+ ion during catalysis (Fig. 5B) -binding sites in proteins (26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37), although the mechanism by which La 3+ influences the enzyme activities of ion-bound complexes has not been studied systematically. Among the lanthanide family, La 3+ exhibits physical and chemical properties similar to Ca 2+ , including a similar ionic radius (1.30 and 1.26 Å, respectively, for eight coordinated metal ions) (38), and acts as a hard Lewis acid that prefers to bind hard bases containing oxygen (39).…”

Effect of Calcium Ions on Enteropeptidase Catalysis

“…It should also be noted that the concentration of lanthanide salts is also much lower than is needed for the detection of the three fold activating effect of Ca 2+ (50 mM). Upon trypsinogen activation by trypsin, lanthanide ions act as activators in a same way as Ca 2+ , but at significantly lower concentrations (1 : 100) [27]. Another unusual attribute of the action of rare earth ions on enteropeptidase is the noncompetitive inhibition mechanism [26].…”

“…Another unusual attribute of the action of rare earth ions on enteropeptidase is the noncompetitive inhibition mechanism [26]. In all these cases of activation of enteropeptidase hydrolysis by calcium ions [1] as well as trypsin hydrolysis by calcium ions [6,7] or lanthanides [27], the increase in the rate of enzymatic reaction occurs mainly due to decrease in the K m value. However, the Schematics of: a) full length bovine enteropeptidase (EP 118 1035); b) autolyzed bovine enteropeptidase (EP 466 1035); c) light chain of recombinant bovine enteropeptidase containing 17 C terminal amino acid residues from the heavy chain (EP 784 1035) a b c addition of lanthanide salts has practically no influence on the K m value for the reaction of trypsinogen activation by enteropeptidase; inhibition is due to decrease in the V max value [26].…”

Non-conventional therapeutics for oral infections

“…71,72 However, lanthanides also activate the conversion of trypsinogen to trypsin. 73 A recently discovered inhibitor of Clp proteases was identified by high throughput screening as N-(1-(2-aminoethyl) -lH-tetrazol-5-yl) -3-chlorobenzamide and which is termed F2. 74 This inhibitor has pleiotropic effects on the bacterial cell because the Clp protease regulates a wide range of genes 75 and also F2 has been shown to increase the effectiveness of bacterial killing by human whole blood, indicating that this compound can augment innate immune defenses.…”