Effect of Zn on acetyl coenzyme a synthase: evidence for a conformational change in the alpha subunit during catalysis.

Acetyl coenzyme A synthase (ACS) is an alpha2beta2 tetramer in which the active-site A-cluster, located in the alpha subunits, consists of an Fe4S4 cubane bridged to a {Nip Nid} binuclear site. The alpha subunits exist in two conformations. In the open conformation, Nip is surface-exposed, while the proximal metal is buried in the closed conformation. Nip is labile and can be replaced by Cu. In this study, the effects of Zn are reported. ACS in which Zn replaced Nip was inactive and did not exhibit the so-called NiFeC EPR signal nor the ability to accept a methyl group from the corrinoid-iron-sulfur protein (CoFeSP). Once Zn-bound, it could not be replaced by subsequently adding Ni. The Zn-bound A-cluster cannot be reduced and bound with CO or become methylated, probably because Zn (like Cu) is insufficiently nucleophilic for these functions. Unexpectedly, Zn replaced Nip only while ACS was engaged in catalysis. Under these conditions, replacement occurred with kapp approximately 0.6 min-1. Replacement was blocked by including EDTA in the assay mix. Zn appears to replace Nip when ACS is in an intermediate state (or states) of catalysis but this(these) state(s) must not be present when ACS is reduced in CO alone, or in the presence of CoA, CoFeSP, or reduced methyl viologen. Nip appears susceptible to Zn-attack when the alpha subunit is in the open conformation and protected from attack when it is in the closed conformation. This is the first evidence that the structurally-characterized conformations of the alpha subunit change during catalysis, indicating a mechanistic role for this conformational change.