Ordered-sequential mechanisms

A substrate analog will frequently inhibit only one of the two forms of a multisubstrate enzyme with a ping-pong mechanism.1 72 Reciprocal plots made for various inhibitor concentrations consist of a family of parallel lines reminiscent of uncompetitive inhibition. Observation of such parallel line plots can support a ping-pong mechanism for an enzyme but cannot prove it because in some cases parallel lines are observed for inhibition of enzymes acting by an ordered sequential mechanism. The following question arises naturally for any ordered bimolecular reaction (Eq. 9-43) Of the... 

Oncogenic viruses 248 One-start helix 334 Oparin, I. V. 9 Open systems 289 Operator sequence of DNA iH NMR spectrum 269 Operons 240 Opsin(s) 553 Optical rotation 42 Optimum rate for enzymes 469 d Orbitals, participation in covalent bond formation 311 Orcinol 251 Ordered binding 464 Ordered sequential mechanism 475 Organelle(s) 11... 

Loewus, M.W., Bedgar, D.L., and Loewus, F.A., 1984, lL-myo-inositol 1-phosphate synthase. An ordered sequential mechanism. J. Biol. Chem. 259 7644-7647. 

In the ordered sequential mechanism, the coenzyme always binds first and the lactate is always released first. This sequence can be represented as follows in a notation developed by W. Wallace Cleland ... 

Many enzymes that have NAD or NADH as a substrate exhibit the ordered sequential mechanism. Consider lactate dehydrogenase, an important enzyme in glucose metabolism (p. 447). This enzyme reduces pyruvate to lactate while oxidizing NADH to NAD. ... 

In an ordered sequential mechanism where C is an alternate substrate for the second substrate, B, one observes competitive inhibition by C versus B and noncompetitive inhibition versus A. for the competitive inhibition is given by an equation analogous to Eq. (25) containing ia, and A in place of ib, b(c). and B. For the noncompetitive inhibition, is given by an equation analogous to Eq. (26), with KaKc K (e), and B replacing K, Ka,... 

Special literature on enzymatic kinetics contains different cases of the ping-pong mechanism as well as an ordered sequential mechanism for bi-substrate and tri-substrate cases which can in fact be rather easily derived from eq. (6.29). 

ATP + 5 -dephospho-DNA = ADP + 5 -phospho-DNA (also acts on 5 -de-phospho-RNA 3 -mononucleotides <8> random reaction mechanism [4] <8> random order sequential mechanism [12] <2, 9> reaction mechanism [21, 27] <2> 5 amino acids, LyslS, Seri 16, Asp35, Arg38, and Argl26, comprise the 5 -kinase active site, structure-activity relationships [25] <2> structure of the active sites for 5 -polynucleotide kinase and 3 phosphatase activity of the bifunctional enzyme, substrate binding mechanism [27])... 

By plotting 1/v versus 1/a at constant b (columns in Table 3.6) values of apparent kinetic parameters Vap and Kap are obtained from the intercepts in the Y and X-axis respectively, as shown in Fig. 3.8 for ordered (I) and random (II) mechanisms. By plotting 1/v versus 1/b at constant a (rows in Table 3.6) values of apparent kinetic parameters Vap and Kap are obtained from the intercepts in the Y and X-axis respectively, as shown in Fig. 3.8 for ordered (I) and random (II) mechanisms. Ordered sequential mechanism can be easily distinguished from random sequential in Lineweaver-Burke plots in the case of ordered mechanism, intercept in the Y-axis (1/Vap ) is a constant (1/V) independent of a (the substrate who binds to the enzyme first), while in the case or random mechanism it depends on a (see Table 3.5). 

Graphical representation of CPBR and CSTR performance is in Fig. 5.6 for one-substrate Michaelis-Menten kinetics and in Figs. 5.7-5.10 for different inhibition kinetics. Graphical representation of CPBR and CSTR performance is presented in Figs. 5.11 and 5.12 for the case of ordered sequential mechanism considering A... 

In the experiment a large excess of thioglycerol was used and thus the two second order reactions can be approximated with a two-step pseudo first order sequential mechanism. Thus, we first attempt to fit this biphasic reaction with a simple consecutive reaction scheme... 

In the ordered-sequential mechanism, there is no formation of a second-substrate complex, as the enzyme initially has no site for binding with the second substrate. This site is formed only when the first substrate has been bound to the enzyme. On the other hand, in the random mechanism, the enzyme can bind with either the first or second substrate to produce the corresponding complex, which can then bind to the other substrate to generate the new complex, which later generates and releases the product. 

Figure 7.2. Fixed substrate concentration dependence for enzymes displaying ordered sequential mechanisms (a) Dependence of on [B] b) dependence of K on [B] (c) dependence of K on [A].

---