Intracellular enzymes can be organized into a variety of assemblies, shuttling intermediates from one active site to the next. Eukaryotic compartmentalization within mitochondria and peroxisomes and substrate tunneling within multi-enzyme complexes have been well recognized. Intriguingly, the central pathways in prokaryotes may also form extensive channels, including the heavily branched glycolysis pathway. In vivo channeling through cascade enzymes is difficult to directly measure, but can be inferred from in vitro tests, reaction thermodynamics, transport/reaction modeling, analysis of molecular diffusion and protein interactions, or steady state/dynamic isotopic labeling. Channeling presents challenges but also opportunities for metabolic engineering applications. It rigidifies fluxes in native pathways by trapping or excluding metabolites for bioconversions, causing substrate catabolite repressions or inferior efficiencies in engineered pathways. Channeling is an overlooked regulatory mechanism used to control flux responses under environmental/genetic perturbations. The heterogeneous distribution of intracellular enzymes also confounds kinetic modeling and multiple-omics analyses. Understanding the scope and mechanisms of channeling in central pathways may improve our interpretation of robust fluxomic topology throughout metabolic networks and lead to better design and engineering of heterologous pathways.

细胞内酶可以组织成各种组装，将中间体从一个活性位点穿梭到另一个活性位点。线粒体和过氧化物酶体中的真核区室化以及多酶复合物中的底物隧穿已得到公认。有趣的是，原核生物中的中央途径也可能形成广泛的通道，包括重度分支的糖酵解途径。通过级联酶的体内通道很难直接测量，但可以从体外推断测试，反应热力学，转运/反应模型，分子扩散和蛋白质相互作用的分析或稳态/动态同位素标记。通道化不仅带来了挑战，而且为代谢工程应用带来了机遇。它通过捕获或排除代谢物进行生物转化来强化天然途径的通量，从而导致底物分解代谢物阻抑或工程途径的效率较低。通道是一种被忽视的调节机制，用于控制环境/遗传扰动下的通量响应。细胞内酶的异质分布也混淆了动力学建模和多组学分析。