Multi-enzyme assemblies

Nature gives us some illustrative examples of iterative methodologies in its biochemical mechanisms. The fatty acid-polyketide biosynthesis is one of them. The assembly of acyl units by sequential Claisen-type condensations to form a polyketide or fatty acid takes place at a multi-enzyme complex, at which the initial molecule is lengthened by one C2-unit per pass of a reaction cycle (Fig. 2). 

Fig. 2.2 Enzyme reactors prepared by LbL assembly (A) reactor on quartz plate for color-indication of glucose (B) multi-enzyme reactor for starch digestion on ultrafilter. Adapted from [26], M. Onda etal, Biotechnol. Bioeng. 1996, 57, 163 and [27], M. Onda et al.,J. Ferment. Bioeng. 1996, 82, 502.

This is not the only example of Nature inventing the assembly line a long time before Henry Ford—both pyru-vate dehydrogenase and a-ketoglutarate dehydrogenase, mentioned earlier in the chapter, are also multi-enzyme complexes. 

Another type of compartmentation involves the assembly of enzymes catalyzing sequential reactions into multi-enzyme complexes so that intermediates of the pathway can be directly transferred from the active site on one enzyme to the active site on another enzyme, thereby preventing loss of energy and information. 

Oyamatsu D, KanayaN, Hirano Y, NishizawaM, Matsue T. Area-selective immobilization of multi enzymes by using the reductive desorption of self-assembled monolayer. Electrochemistry 2003 71 439-441. 

Given that these proteins have properly assembled, the initiation complex is ready to start transcription. How does the enzyme get started A component of TFIID, again a multi-subunit complex TFIIH, unwinds the DNA and phosphorylates serine-5 of the C-terminal tail (CTD) of the largest polymerase subunit (Rpbl). Serine-5 phosphorylation and phosphorylation of serine-2 (by pTEFb) are required to release the enzyme from the other components of the initiation complex and to start RNA synthesis. 

It is noted that the anodic peak current prominently increases with an increase in the molar ratio of ferrocene to glucose oxidase whilst the amount of enzyme self-assembled on the electrode surface is fixed as presented in Figs. 14-16. This indicates that each modified ferrocene may contribute to electron transfer between the enzyme and the electrode in the case of gold-black electrode, the ferrocene-modified enzyme could form multi electron transfer paths on the porous gold-black electrode. 

Mother nature has resolved the various limitations involved in multi-electron processes. Unique assemblies composed of cofactors and enzymes provide the microscopic catalytic environments capable of activating the substrates, acting as multi-electron relay systems and inducing selectivity and specificity. Artificially tailored heterogeneous and homogeneous catalysts as well as biocatalysts (enzymes and cofactors) are, thus, essential ingredients of artificial photosynthetic devices. 

Little is known of the ways of biosynthesis and insertion in the membranes of PolyP-PHB-Ca2+ complexes. In polyphosphate kinase 1 mutants of E. coli, the amounts of the complexes did not change (Castuma et al., 1995). Therefore, the PolyP in the complexes is synthesized not by polyphosphate kinase 1 but by another enzyme. E. coli strain, which lacks the AcrA component of a major multi-drug resistance pump, had greatly reduced amounts of the complexes and was defective in its ability to maintain sub-micromolar levels of free Ca2+ in the cytoplasm (Jones et al., 2003). This indicates that the AcrAB transporter may have a novel, hitherto undetected, physiological role, either directly in the membrane assembly of the PHB complexes or the transport of a component of the membrane, which is essential for assembly of the complexes into the membrane. 

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