The origins of enzyme catalysis have been attributed to both transition-state stabilization as well as ground-state destabilization of the substrate. For the latter paradigm, the enzyme orotidine-5′-monophosphate decarboxylase (OMPDC) serves as a reference system as it contains a negatively charged residue at the active site that is thought to facilitate catalysis by exerting an electrostatic stress on the substrate carboxylate leaving group. Snapshots of how the substrate binds to the active site and interacts with the negative charge have remained elusive. Here we present crystallographic snapshots of human OMPDC in complex with the substrate, substrate analogues, transition-state analogues and product that defy the proposed ground-state destabilization by revealing that the substrate carboxylate is protonated and forms a favourable low-barrier hydrogen bond with a negatively charged residue. The catalytic prowess of OMPDC almost entirely results from the transition-state stabilization by electrostatic interactions of the enzyme with charges spread over the substrate. Our findings bear relevance for the design of (de)carboxylase catalysts.

酶催化的起源归因于底物的过渡态稳定化和基态去稳定化。对于后一种范例，酶 orotidine-5'-monophosphate decarboxylase (OMPDC) 用作参考系统，因为它在活性位点含有带负电荷的残基，被认为通过在底物羧酸离去基团上施加静电应力来促进催化. 关于底物如何与活性位点结合并与负电荷相互作用的快照仍然难以捉摸。在这里，我们展示了与底物、底物类似物复合的人类 OMPDC 的晶体学快照，过渡态类似物和产物通过揭示底物羧酸盐被质子化并与带负电荷的残基形成有利的低势垒氢键来挑战所提出的基态去稳定化。OMPDC 的催化能力几乎完全来自于通过酶与散布在底物上的电荷的静电相互作用实现的过渡态稳定化。我们的研究结果与（脱）羧化酶催化剂的设计相关。