Cleavage of farnesylated COOH-terminal heptapeptide of mouse N-ras by brain microsomal membranes: Evidence for a carboxypeptidase which specifically removes the COOH-terminal methionine

Akopyan, T. N.; Couedel, Y.; Beaumont, Ann; Fournié‐Zaluski, Marie‐Claude; Roques, B. P.

doi:10.1016/0006-291x(92)90449-u

Cited by 10 publications

(12 citation statements)

References 15 publications

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“…At pH 7.6, but not at pH 9.5, [3H]S is produced by direct proteolysis of ECB-C(F)VI[3H]S. This type of carboxypeptidase activity was not seen in the studies with bovine liver microsomes (Ma & Rando, 1992), but Akopyan et al (1992) reported that incubation of the prenylated peptide propionyl-GSOC(F)VLM with mouse brain microsomes results in the release of methionine as the major product along with small amounts of leucine. Interestingly, the pH optimum for this mouse brain enzyme is around 7.2, which is similar to our findings for the production of [3H]S from ECB-C(F) VI[3H]S.…”

Section: Discussionmentioning

confidence: 90%

“…The major product formed is the tripeptide, Ali-Ali-Xaa, along with smaller amounts of dipeptide and single amino acid. Akopyan and co-workers observed that incubation of the prenylated peptide propionyl-GSPC(F) VLM [S-farnesylated and 5-geranylgeranylated cysteine residues are designated as C(F) and C(GG), respectively] with mouse brain microsomes results in the release of methionine as the major product along with small amounts of leucine (Akopyan et al, 1992). Hrycyna and Clarke reported that yeast membranes are capable of cleaving the synthetic peptide JV-acetyl-KSKTKC-(F)VIM to generate the substrate for the prenyl-cysteine -carboxylmethyltransferase [TV-acetyl-KSKTKC(F)] (Hrycyna & Clarke, 1992) but the identity of the C-terminal peptide product(s) was not determined.…”

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confidence: 99%

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A prenylated protein-specific endoprotease in rat liver microsomes that produces a carboxyl-terminal tripeptide

Jang¹,

Yokoyama²,

Gelb³

1993

Biochemistry

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The maturation of proteins that contain the C-terminal sequence Cys-Ali-Ali-Xaa (where Ali is usually an aliphatic amino acid and Xaa is a number of different amino acids) involves the attachment of a farnesyl or geranylgeranyl group to the cysteine residue, proteolytic removal of the C-terminal three amino acids, and methylation of the prenylated cysteine alpha-carboxyl group. Two prenylated and radiolabeled peptides were prepared in order to detect the proteolysis step(s) in a cell-free system and to determine the reaction products. These peptides are ECB-NPFRQRRFFC(S-geranylgeranyl)AI[3H]L and ECB-C(S-farnesyl)VI[3H]S (ECB is an extended-chain biotin group) which are patterned after the C-termini of geranylgeranylated and farnesylated G protein gamma-subunits. Incubation of these peptides with rat liver microsomes, but not cytosol, results in the production of radiolabeled dipeptides (I[3H]L and I[3H]S) and tripeptides (AI[3H]L and VI[3H]S) as the major products and smaller amounts of amino acids ([3H]L and [3H]S). A multitude of independent experiments shows that the dipeptides are produced from the tripeptides by secondary proteolysis. Although a portion of the [3H]S produced comes directly from ECB-C(S-farnesyl)VI[3H]S, the KM of > 30 microM for this reaction is significantly higher than the KM of 1.1 microM for the production of VI[3H]S from the farnesylated peptide. This suggests that the carboxypeptidase is not part of the pathway for the maturation of prenylated proteins. Nonprenylated peptides at concentrations of 10-100-fold higher than those of the prenylated substrates did not reduce the amount of tripeptide produced.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Discussionmentioning

confidence: 90%

mentioning

confidence: 99%

A prenylated protein-specific endoprotease in rat liver microsomes that produces a carboxyl-terminal tripeptide

Jang¹,

Yokoyama²,

Gelb³

1993

Biochemistry

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“…Our observation that substrate context impacts the effectiveness of Rce1p inhibitors could explain the inconsistent effect of serine protease inhibitors and alkylating agents on Rce1p activity [24,27,30,31]. We thus suggest that evaluating inhibitors of Rce1p in the context of multiple substrates is the best way to fully ascertain the inhibitory properties of a particular compound for this multi-substrate enzyme.…”

Section: Discussionmentioning

confidence: 92%

“…Several inhibitors of Rce1p in addition to those discussed above have been described [23][24][25][26][27][30][31][32][33][34][35]. These fall into two classes: general non-specific inhibitors and substrate mimetics.…”

Section: Introductionmentioning

confidence: 99%

“…Rce1p is reportedly insensitive to many broadspectrum inhibitors, such as EDTA, EGTA, antipain, chymostatin, pepstatin A, leupeptin, E64, and is partially sensitive to MMTS. It should be noted that the sensitivity of Rce1p to the serine protease inhibitors PMSF and DFP and the alkylating agent NEM has been inconsistently reported [24,27,30,31]. The second class of Rce1p inhibitors is represented by substrates with non-cleavable peptide bonds, isoprenoid-like compounds, and bisubstrate analogues (i.e., compounds containing both a farnesylmimetic and peptidomimetic).…”

Section: Introductionmentioning

confidence: 99%

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Inhibition of the CaaX proteases Rce1p and Ste24p by peptidyl (acyloxy)methyl ketones

Porter

Hildebrandt

Breevoort

et al. 2007

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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The CaaX proteases Rce1p and Ste24p can independently promote a proteolytic step required for the maturation of certain isoprenylated proteins. Although functionally related, Rce1p and Ste24p are unrelated in primary sequence. They have distinct enzymatic properties, which are reflected in part by their distinct inhibitor profiles. Moreover, Rce1p has an undefined catalytic mechanism, whereas Ste24p is an established zinc-dependent metalloprotease. This study demonstrates that both enzymes are inhibited by peptidyl (acyloxy)methyl ketones (AOMKs), making these compounds the first documented dual specificity inhibitors of the CaaX proteases. Further investigation of AOMK-mediated inhibition reveals that varying the peptidyl moiety can significantly alter the inhibitory properties of AOMKs toward Rce1p and Ste24p and that these enzymes display subtle differences in sensitivity to AOMKs. This observation suggests that this compound class could potentially be engineered to be selective for either of the CaaX proteases. We also demonstrate that the reported sensitivity of Rce1p to TPCK is substrate-dependent, which significantly alters the interpretation of certain reports having used TPCK sensitivity for mechanistic classification of Rce1p. Finally, we show that an AOMK inhibits the isoprenylcysteine carboxyl methyltransferase Ste14p. In sum, our observations raise important considerations regarding the specificity of agents targeting enzymes involved in the maturation of isoprenylated proteins, some of which are being developed as anti-cancer therapeutic agents.

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A Case for ras Targeted Agents as Antineoplastics

Sebolt–Leopold¹

1997

Cancer Therapeutics

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Cleavage of farnesylated COOH-terminal heptapeptide of mouse N-ras by brain microsomal membranes: Evidence for a carboxypeptidase which specifically removes the COOH-terminal methionine

Cited by 10 publications

References 15 publications

A prenylated protein-specific endoprotease in rat liver microsomes that produces a carboxyl-terminal tripeptide

A prenylated protein-specific endoprotease in rat liver microsomes that produces a carboxyl-terminal tripeptide

Inhibition of the CaaX proteases Rce1p and Ste24p by peptidyl (acyloxy)methyl ketones

A Case for ras Targeted Agents as Antineoplastics

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