1wkm
From Proteopedia
|
THE PRODUCT BOUND FORM OF THE MN(II)LOADED METHIONINE AMINOPEPTIDASE FROM HYPERTHERMOPHILE PYROCOCCUS FURIOSUS
Overview
Methionine aminopeptidases (MetAPs) are ubiquitous metallohydrolases that, remove the N-terminal methionine from nascent polypeptide chains. Although, various crystal structures of MetAP in the presence of inhibitors have, been solved, the structural aspects of the product-bound step has received, little attention. Both perpendicular- and parallel-mode electron, paramagnetic resonance (EPR) spectra were recorded for the Mn(II)-loaded, forms of the type-I (Escherichia coli) and type-II (Pyrococcus furiosus), MetAPs in the presence of the reaction product l-methionine (L-Met). In, general, similar EPR features were observed for both, [MnMn(EcMetAP-I)]-L-Met and [MnMn(PfMetAP-II)]-L-Met. The observed, perpendicular-mode EPR spectra consisted of a six-line hyperfine pattern, at g = 2.03 (A = 8.8 mT) with less intense signals with eleven-line, splitting at g = 2.4 and 1.7 (A = 4.4 mT). The former feature results from, mononuclear, magnetically isolated Mn(II) ions and this signal are 3-fold, more intense in the [MnMn(PfMetAP-II)]-L-Met EPR spectrum than in the, [MnMn(EcMetAP-I)]-L-Met spectrum. Inspection of the EPR spectra of both, [MnMn(EcMetAP-I)]-L-Met and [MnMn(PfMetAP-II)]-L-Met at 40 K in the, parallel mode reveals that the [Mn(EcMetAP-I)]-L-Met spectrum exhibits a, well-resolved hyperfine split pattern at g = 7.6 with a hyperfine, splitting constant of A = 4.4 mT. These data suggest the presence of a, magnetically coupled dinuclear Mn(II) center. On the other hand, a similar, feature was not observed for the [MnMn(PfMetAP-II)]-L-Met complex., Therefore, the EPR data suggest that L-Met binds to [MnMn(EcMetAP-I)], differently than [MnMn(PfMetAP-II)]. To confirm these data, the X-ray, crystal structure of [MnMn(PfMetAP-II)]-L-Met was solved to 2.3 A, resolution. Both Mn1 and Mn2 reside in a distorted trigonal bipyramidal, geometry, but the bridging water molecule, observed in the, [CoCo(PfMetAP-II)] structure, is absent. Therefore, L-Met binding, displaces this water molecule, but the carboxylate oxygen atom of L-Met, does not bridge between the two Mn(II) ions. Instead, a single carboxylate, oxygen atom of L-Met interacts with only Mn1, while the N-terminal amine, nitrogen atom binds to M2. This L-Met binding mode is different from that, observed for L-Met binding [CoCo(EcMetAP-I)]. Therefore, the catalytic, mechanisms of type-I MetAPs may differ somewhat from type-II enzymes when, a dinuclear metalloactive site is present.
About this Structure
1WKM is a Single protein structure of sequence from Pyrococcus furiosus with MN and MET as ligands. Active as Methionyl aminopeptidase, with EC number 3.4.11.18 Full crystallographic information is available from OCA.
Reference
EPR and X-ray crystallographic characterization of the product-bound form of the MnII-loaded methionyl aminopeptidase from Pyrococcus furiosus., Copik AJ, Nocek BP, Swierczek SI, Ruebush S, Jang SB, Meng L, D'souza VM, Peters JW, Bennett B, Holz RC, Biochemistry. 2005 Jan 11;44(1):121-9. PMID:15628852
Page seeded by OCA on Wed Nov 21 05:29:37 2007
