Mapping the conformational space accessible to catechol-O-methyltransferase

Ehler, A.; Benz, J.; Schlatter, D.; Rudolph, M.G.

doi:10.1107/S1399004714012917

Figure 6
Conformational changes in the Mg²⁺-binding site. This figure recapitulates some of the information in Fig. 5

from a different angle while focusing on the Mg²⁺ site. (a) and (b) show the closed state of COMT with an octahedral coordination of the Mg²⁺ ion (green sphere). Lys187 is in the in-conformation. (c)–(f) show the half-closed conformation with or without a ligand (SAH or sinefungin) in the adenine site. Lys187 is in the out-conformation. (g) and (h) are the open conformations where neither substrate-binding site is formed. Lys187 is in an intermediate conformation. (a) Structure (8) with sinefungin and tolcapone represents the Michaelis complex with SAM replaced by the inhibitor sinefungin and the substrate catechol replaced by the inhibitor tolcapone. (b) (11b) has Mg²⁺ bound but instead of a catechol two water molecules complete the coordination sphere of Mg²⁺. (c) The sinefungin-bound structure (7) lacks the Mg²⁺ ion but has a K⁺ ion bound to a nearby site. K⁺ coordinates the former Mg²⁺-binding side chains Asp184 and Asn213. The primary ammonium group of sinefungin bridges via a water molecule to the K⁺ ion. Lys89, which normally neutralizes the charge of Asp212, reorients towards the carboxylate of SAH. (d) Structure (10) bound to SAH recapitulates all of the salient features of (c). (e) (11a) bound to SAH has no K⁺ ion bound but retains most of the side-chain positions with the exception of Asn84 in the α2/α3 loop, which moves to contact a water molecule situated at the former position of the Mg²⁺ ion. (f) Structure (9) is interesting as it adopts the same half-closed conformation as (7), (10) and (11a) in the absence of a ligand in the adenine site. As in (e), water replaces Mg²⁺. Sulfate is located at the site where the carboxylate of SAM and related compounds normally binds. In the absence of a ribose, Glu133 reorients to hydrogen-bond to the main-chain amide N atom of Ala110. (g) The apo COMT structure (6) also does not have a functional Mg²⁺ site and water hydrogen-bonds to Asp184 and Asp212. Pro217 in the β5/α9 loop, which normally engages in van der Waals contact with the aryl part of the catechol substrate, has maximally swung away from the body of COMT. (h) Structure (5) has the same overall conformation as structure (6) in (g).

BIOLOGICAL
CRYSTALLOGRAPHY

ISSN: 1399-0047

Volume 70| Part 8| July 2014| Pages 2163-2174

doi:10.1107/S1399004714012917

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