Multienzyme complex, substrate channeling

While PAL itself is recovered from the soluble fractions of cellular lysates via ultracentrifugation purifications, strong experimental evidence suggests it exists as part of a multienzyme complex associated with the endoplasmic reticulum (ER), forming a metabolic channel tunneling substrates directly from one enzymatic reactive site to another (Winkel 2004). The enzyme physically anchored to the ER is cinnamate... 

Substrate Channeling through Multienzyme Complexes May Occur in the Citric Acid Cycle... 

Review of the roles of swinging arms containing lipoate, biotin, and pantothenate in substrate channeling through multienzyme complexes. 

The last three steps of this four-step sequence are catalyzed by either of two sets of enzymes, with the enzymes employed depending on the length of the fatty acyl chain. For fatty acyl chains of 12 or more carbons, the reactions are catalyzed by a multienzyme complex associated with the inner mitochondrial membrane, the trifunctional protein (TFP). TFP is a heterooctamer of 4/34 subunits. Each a subunit contains two activities, the enoyl-CoA hydratase and the /3-hydroxyacyl-CoA dehydrogenase the /3 subunits contain the thiolase activity. This tight association of three enzymes may allow efficient substrate channeling from one active site to the... 

The limit imposed by the rate of diffusion in solution can also be partly overcome by confining substrates and products in the limited volume of a multienzyme complex. Indeed, some series of enzymes are associated into organized assemblies (Section 17.1.9) so that the product of one enzyme is very rapidly found by the next enzyme. In effect, products are channeled from one enzyme to the next, much as in an assembly line. 

The efficiency of the citric acid cycle may be enhanced by the arrangement of the constituent enzymes. Evidence is accumulating that the enzymes are physically associated with one another to facilitate substrate channeling between active sites. The word metabolon has been suggested as the name for such multienzyme complexes. 

In the cell, compartmentation of enzymes into multienzyme complexes or organelles provides a means of regulation, either because the compartment provides unique conditions or because it limits or channels access of the enzymes to substrates. Enzymes or pathways with a common function are often assembled into organelles. For example, enzymes of the TCA cycle are all located within the mitochondrion. The enzymes catalyze sequential reactions, and the product of one reaction is the substrate for the next reaction. The concentration of the pathway intermediates remains much higher within the mitochondrion than in the surrounding cellular cytoplasm. 

The primary sequence of the 0-subunit (397 residues) is also known, and extensive studies have been reported on the 3-dimensional structure of the 0202 multienzyme complex, in particular for the enzyme from Salmonella lyphimurium. The enzymes from Ecoli and S. lyphimurium are very similar the respective a-subunits both consist 268 amino acid residues and differ at 40 positions (15%), while the 0-monomers display only 3.5% difference in primary sequence (both consist of 397 residues, and only 14 of these are different). Crystallographic studies show that the active sites of the a- and 0-subunits are 25 A apart and connected by a tunnel, which presumably serves to carry the one metabolic intermediate (indole) from the active site of the a-subunit to the active site of the p-subunit. The kinetics of this substrate channeling have been studied by chemical quench-flow and stopped-flow methods- (I.P. Crawford J.Ito Proc. NatL Acad. ScL USA. 51 (1964) 390-397 B.P.Nichols CYanofeky Proc NatL Acad. Sci. USA. 76 (1979) 5244-5248 S-A. Ahmed et al. J. Biol. Chan. 260 (1985) 3716-3718 CCHyde et al. J. Biol. Chem. 263 (1988) 17857-17871 K.S. Anderson et al. J. Biol. Chem. 266 (1991) 8020-8033]... 

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