Aminopeptidase, methionine

Systematic screening experiments have identified more than 100 synthetic compounds with potent antiangiogenic activity. The mode of action for most of these molecules is not well understood, but some of the 40 compounds are well advanced in clinical trials (Table 3). The first substance to have entered clinical trials was the Fumagillin-derivative AGM 1470. Fumagillin is an antibiotic which inhibits bFGF- and PDGF-induced endothelial cell proliferation. The mechanism of action of AGM 1470 is poorly understood, but it was shown that it binds and inhibits the metalloprotease methionine aminopeptidase (MetAp-2). 

Non-corrin cobalt has a number of interesting applications in the chemical industry, for example in the hydroformylation (OXO) reaction between CO, H2 and olefins. A number of non-corrin Co-containing enzymes have been described, including methionine aminopep-tidase, prolidase, nitrile hydratase and glucose isomerase. We describe the best characterized of these, namely the E. coli methionine aminopeptidase, a ubiquitous enzyme, which cleaves N-terminal methionine from newly translated polypeptide chains. The active site of the enzyme (Figure 15.13) contains two Co(II) ions that are coordinated by the side-chain atoms of five amino acid residues. The distance between the two Co2+ is similar to that between the two Zn2+ atoms in leucine aminopeptidase, and indeed the catalytic mechanism of methionine aminopeptidase shares many features with other metalloproteases, in particular leucine aminopeptidases. 

Figure 15.13 Structure of the bimetallic core of K coli methionine aminopeptidase. (From Kobayashi and Shimizu, 1999. Reproduced with permission of Blackwell Publishing Ltd.)...

Liu S, Widom J, Kemp CW, Crews CM, Clardy J. (1998) Structure of human methionine aminopeptidase-2 complexed with fumagillin. Science 282 1324-1327. 

Sin N, Meng L, Wang MQW, Wen JJ, Bommann WG Crews CM. (1997) The anti-angiogenic agent fumagillin covalently binds and inhibits the methionine aminopeptidase, MetAP-2. Proc Natl Acad Sci USA 94 6099-6103. 

Many aminopeptidases are metalloenzymes.437 Most studied is the cytosolic leucine aminopeptidase which acts rapidly on N-terminal leucine and removes other amino acids more slowly. Each of the subunits of the hexameric enzyme contains two divalent metal ions, one of which must be Zn2+ or Co2+438/439 A methionine aminopeptidase from E. colt contains two Co2+ ions440/441 and a proline-specific aminopeptidase from the same bacterium two Mn2+.442 In all of these enzymes the metal ions are present as dimetal pairs similar to those observed in phosphatases and discussed in Section D,4 and to the Fe-Fe pairs of hemerythrin and other diiron proteins (Fig. 16-20). A hydroxide ion that bridges the metal ions may serve as the nucleophile in the aminopeptidases.438 A bound bicarbonate ion may assist.4383... 

Endo H, Takenaga K, Kanno T, Satoh H, Mori S. 2002. Methionine aminopeptidase 2 is a new target for the metastasis-associated protein, S100A4. I Biol Chem 277(29) 26396-26402. 

PJKeeling, WF Doolittle. Methionine aminopeptidase-1 the MAP ofthe mitochondrion Trends Biochem Sci 21 285-286, 1996. 

SY Chang, EC McGary, S Chang. Methionine aminopeptidase gene of Escherichia coli is essential for cell growth. J Bacteriol 171 4071-4072, 1989. 

Lancelot C, Mathot S, Owens NJP (1986) Modelling protein synthesis, a step to an accurate estimate of net primary production the case of Phaeocystis pouchetii colonies in Belgian coastal waters. Mar Ecol Prog Ser 32 193-202 Lancelot C, Billen G, Sournia A, Weisse T, Colijin F, Veldhuis MJW, Davies A, Wassman P (1987) Phaeocystis blooms and nutrient enrichment in the continental coastal zones of the North Sea. AMBIO 16(1) 38 16 Lewandowska J, Kosakowska A (2004) Effect of iron limitation on cells of the diatom Cyclotella meneghiniana Kutzing. Oceanologia 46(2) 269-287 Lowther WT, Matthews BW (2000) Structure and function of the methionine aminopeptidases. Biochim Biophys Acta 1477 157-167... 

The /V-formylmethionine of a nascent protein synthesized in bacteria is removed by the sequential activities of PDF and a methionine aminopeptidase to generate the mature protein. The gene encoding PDF was cloned and overexpressed in E. coli by Meinnel and coworkers (1993). The PDF enzyme has an unusual metal ion (Fe2+) as its catalyst. However, the ferrous ion in this enzyme is unstable and can be quickly and irreversibly oxidized to ferric ion, rapidly inactivating the enzyme. PDF-based assay development therefore depended on the ability of nickel ion to replace ferrous ion in vitro, increasing the stability of the enzyme and maintaining its enzymatic activity (Groche et al., 1998 Clements et al., 2001 Hackbarth et al., 2002). 

Many crystal structures of other potassium-activated enzymes have been reported. They generally require a divalent cation and are activated by a monovalent cation that also binds. Such enzymes include amylase, which has a unique Ca Na a trinuclear center, 5 -adenosyImethionine synthetase, E. coli methionine aminopeptidase in which... 

Virtually all proteins expressed in cells undergo processing to remove the N-terminal methionine residue encoded by the start codon , ATG. Cleavage of this initiation methionine from newly formed polypeptide chains is catalyzed by the enzyme methionine aminopeptidase (equation 9). 

The active site of methionine aminopeptidase contains a binuclear cobalt complex that is required for activity, although a number of divalent metal ions support turnover to varying degrees. X-ray crystallographic studies on the enzyme in complexes with transition state analogs suggests that the binuclear metal cluster serves to stabilize the tetrahedral intermediate in peptide hydrolysis. ... 

Lowther, W.T., and Matthews, B.W., Strnctnre and function of the methionine aminopeptidases, Biochim. Biophys. Acta 1477, 157-167, 2000. 

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