Structural homology among mammalian and Saccharomyces cerevisiae isoprenyl-protein transferases.

Kohl, Nancy E.; Diehl, Ronald E.; Schaber, Michael D.; Rands, Elaine; Soderman, Denis D.; He, Bin; Moores, Sheri L.; Pompliano, D L; Ferro‐Novick, Susan; Powers, S

doi:10.1016/s0021-9258(18)55146-2

Cited by 91 publications

(30 citation statements)

References 23 publications

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“…These enzymes include ( a ) the farnesyl transferase complex (Ram1p/Ram2p); ( b ) the CAAX prenyl proteases that carry out AAXing (Rce1p or Ste24p [see below]); and ( c ) the carboxyl methyltransferase (Ste14p) (Hrycyna and Clarke, 1990;Schafer et al, 1990;He et al, 1991;Boyartchuk et al, 1997). The genes encoding mammalian CAAX processing enzymes that have been cloned to date are the rat, bovine, and human farnesyltransferases; the rat and human geranylgeranyltransferase; and the human prenyl protein carboxylmethyltransferase (Kohl et al, 1991;Andres et al, 1993;Zhang, 1994 [no. 1610]; Dai et al, 1998). In contrast, no mammalian CAAX protease genes have as yet been identified.…”

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confidence: 99%

Dual Roles for Ste24p in Yeast a-Factor Maturation: NH2-terminal Proteolysis and COOH-terminal CAAX Processing

Tam

Nouvet

Fujimura-Kamada

et al. 1998

The Journal of Cell Biology

125

192

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Maturation of the Saccharomyces cerevisiae a-factor precursor involves COOH-terminal CAAX processing (prenylation, AAX tripeptide proteolysis, and carboxyl methylation) followed by cleavage of an NH2-terminal extension (two sequential proteolytic processing steps). The aim of this study is to clarify the precise role of Ste24p, a membrane-spanning zinc metalloprotease, in the proteolytic processing of the a-factor precursor. We demonstrated previously that Ste24p is necessary for the first NH2-terminal processing step by analysis of radiolabeled a-factor intermediates in vivo (Fujimura-Kamada, K., F.J. Nouvet, and S. Michaelis. 1997. J. Cell Biol. 136:271–285). In contrast, using an in vitro protease assay, others showed that Ste24p (Afc1p) and another gene product, Rce1p, share partial overlapping function as COOH-terminal CAAX proteases (Boyartchuk, V.L., M.N. Ashby, and J. Rine. 1997. Science. 275:1796–1800). Here we resolve these apparently conflicting results and provide compelling in vivo evidence that Ste24p indeed functions at two steps of a-factor maturation using two methods. First, direct analysis of a-factor biosynthetic intermediates in the double mutant (ste24Δ rce1Δ) reveals a previously undetected species (P0*) that fails to be COOH terminally processed, consistent with redundant roles for Ste24p and Rce1p in COOH-terminal CAAX processing. Whereas a-factor maturation appears relatively normal in the rce1Δ single mutant, the ste24Δ single mutant accumulates an intermediate that is correctly COOH terminally processed but is defective in cleavage of the NH2-terminal extension, demonstrating that Ste24p is also involved in NH2-terminal processing. Together, these data indicate dual roles for Ste24p and a single role for Rce1p in a-factor processing. Second, by using a novel set of ubiquitin–a-factor fusions to separate the NH2- and COOH-terminal processing events of a-factor maturation, we provide independent evidence for the dual roles of Ste24p. We also report here the isolation of the human (Hs) Ste24p homologue, representing the first human CAAX protease to be cloned. We show that Hs Ste24p complements the mating defect of the yeast double mutant (ste24Δ rce1Δ) strain, implying that like yeast Ste24p, Hs Ste24p can mediate multiple types of proteolytic events.

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Dual Roles for Ste24p in Yeast a-Factor Maturation: NH2-terminal Proteolysis and COOH-terminal CAAX Processing

Tam

Nouvet

Fujimura-Kamada

et al. 1998

The Journal of Cell Biology

125

192

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“…Short peptides encompassing the CaaX motif of these substrates are also recognized by the enzyme (Reiss et al, 1990(Reiss et al, , 1991b. Native FPT has been purified from rat and bovine brain cytosol and is a heterodimeric protein of ∼95 kDa containing two subunits designated as R (48 kDa) and β (45 kDa) (Reiss et al, 1990;Kohl et al, 1991). cDNA clones for both subunits have been isolated from rat, bovine, and human cDNA libraries (Chen et al, 1991a,b;Omer et al, 1993).…”

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Substitution of Cadmium for Zinc in Farnesyl:Protein Transferase Alters Its Substrate Specificity

Casey

et al. 1996

Biochemistry

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Ras proteins are mutationally activated in a variety of human cancers. Since farnesylation of Ras proteins is required for expression of their oncogenic potential, the enzyme responsible for this reaction, farnesyl:protein transferase (FPT), has become a major target for anticancer drug development. FPT is a zinc metalloenzyme, and the zinc is essential for its catalytic activity. To begin to elucidate the role of zinc in catalysis, we initiated metal substitution studies. Of all metals tested, only cadmium was able to functionally substitute for zinc, reconstituting enzymatic activity with native substrates (H-Ras and farnesyl diphosphate) to about 50% of that of the zinc-containing enzyme. Several important differences were observed between cadmium-substituted FPT (Cd-FPT) and zinc-containing FPT (Zn-FPT). Cd-FPT not only uses H-ras with its native CaaX motif (Ras-CVLS) as a substrate but also can farnesylate H-ras in which the CaaX motif is altered to contain a C-terminal leucine residue (Ras-CVLL). Similarly, Cd-FPT can farnesylate leucine-terminated peptides. Leucine-terminated proteins and peptides are usually substrates for the related enzyme geranylgeranyl:protein transferase type I. Farnesylation of Ras-CVLS and Ras-CVLL by Cd-FPT exhibited similar sensitivity to the FPT inhibitor SCH 44342 and to the peptide inhibitor CAIM. However, unlike Zn-FPT, Cd-FPT is also potently inhibited by the leucine-terminated peptide CAIL. These results indicate that the metal ion content of FPT strongly influences its protein substrate specificity.

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“…Protein farnesylation and geranylgeranylation are catalyzed by distinct prenyltransferases, and the type of prenyl group that is added to the protein is dictated primarily by the C-terminal amino acid (Xaa) (Casey et al, 1991;Finegold et al, 1991;Kohl et al, 1991;Moores et al, 1991;Seabraetal., 1991;Yokoyama etal., 1991;Yoshida et al, 1991). The prenyl-cysteine a-carboxylmethyltransferase that utilizes S-adenosylmethionine as the methyl donor has been detected in the membrane fraction of yeast cells and mammalian tissue (Perez-Sala et al, 1991; Stephenson & .…”

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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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Structural homology among mammalian and Saccharomyces cerevisiae isoprenyl-protein transferases.

Cited by 91 publications

References 23 publications

Dual Roles for Ste24p in Yeast a-Factor Maturation: NH2-terminal Proteolysis and COOH-terminal CAAX Processing

Dual Roles for Ste24p in Yeast a-Factor Maturation: NH2-terminal Proteolysis and COOH-terminal CAAX Processing

Substitution of Cadmium for Zinc in Farnesyl:Protein Transferase Alters Its Substrate Specificity

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

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