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THE PRODUCT BOUND FORM OF THE MN(II)LOADED METHIONINE AMINOPEPTIDASE FROM HYPERTHERMOPHILE PYROCOCCUS FURIOSUSTHE PRODUCT BOUND FORM OF THE MN(II)LOADED METHIONINE AMINOPEPTIDASE FROM HYPERTHERMOPHILE PYROCOCCUS FURIOSUS
Structural highlights
FunctionMAP2_PYRFU Removes the N-terminal methionine from nascent proteins. The N-terminal methionine is often cleaved when the second residue in the primary sequence is small and uncharged (Met-Ala-, Cys, Gly, Pro, Ser, Thr, or Val).[HAMAP-Rule:MF_01975][1] Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedMethionine 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. 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[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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