Polylysine and CVIM sequences of K-RasB dictate specificity of prenylation and confer resistance to benzodiazepine peptidomimetic in vitro

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Abstract

BZA-5B, a benzodiazepine peptidomimetic, inhibits CAAX farnesyltransferase (FTase) and blocks attachment of farnesyl groups to oncogenic and wild-type H-Ras in animal cells. This compound slows the growth of cells transformed with oncogenic H-Ras at concentrations that do not affect the growth of nontransformed cells. This finding suggested that nontransformed cells may produce a form of Ras whose prenylation is resistant to BZA-5B. In the current studies, we found that FTase had a 50-fold higher affinity for K- RasB than for H-Ras in vitro. Farnesylation of K-RasB was inhibited by BZA- 2B, the active form of BZA-5B, but only at concentrations that were 8-fold higher than those that inhibited farnesylation of H-Ras. K-RasB, but not H- Ras, was also a substrate for CAAX geranylgeranyltransferase-1 (GGTase-1), and its affinity for the enzyme was equal to that of Rap1B, an authentic leucine-terminated substrate for GGTase-1. Inhibition of the geranylgeranylation of K-RasB occurred only at high concentrations of BZA- 2B. All of these properties of K-RasB were traced to the combined effects of its COOH-terminal CVIM sequence and the adjacent polylysine sequence, neither of which is present in H-Ras. These studies provide a potential explanation for the resistance of nontransformed cells to growth inhibition by BZA-5B. Inasmuch as the majority of Ras-related human cancers contain oncogenic versions of K-RasB rather than H-Ras, the current data suggest that in vitro studies of FTase inhibitors with potential anti-cancer activity should use authentic K-RasB as a substrate.

Original languageEnglish (US)
Pages (from-to)6221-6226
Number of pages6
JournalJournal of Biological Chemistry
Volume270
Issue number11
DOIs
StatePublished - 1995

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Peptidomimetics
Prenylation
Polylysine
Benzodiazepines
Farnesyltranstransferase
Cells
Substrates
Growth
Leucine
Animals
Neoplasms
In Vitro Techniques
BZA 5B
Enzymes
BZA-2B

ASJC Scopus subject areas

  • Biochemistry

Cite this

@article{e192bdf3a19d45808d41b776c6dc095a,
title = "Polylysine and CVIM sequences of K-RasB dictate specificity of prenylation and confer resistance to benzodiazepine peptidomimetic in vitro",
abstract = "BZA-5B, a benzodiazepine peptidomimetic, inhibits CAAX farnesyltransferase (FTase) and blocks attachment of farnesyl groups to oncogenic and wild-type H-Ras in animal cells. This compound slows the growth of cells transformed with oncogenic H-Ras at concentrations that do not affect the growth of nontransformed cells. This finding suggested that nontransformed cells may produce a form of Ras whose prenylation is resistant to BZA-5B. In the current studies, we found that FTase had a 50-fold higher affinity for K- RasB than for H-Ras in vitro. Farnesylation of K-RasB was inhibited by BZA- 2B, the active form of BZA-5B, but only at concentrations that were 8-fold higher than those that inhibited farnesylation of H-Ras. K-RasB, but not H- Ras, was also a substrate for CAAX geranylgeranyltransferase-1 (GGTase-1), and its affinity for the enzyme was equal to that of Rap1B, an authentic leucine-terminated substrate for GGTase-1. Inhibition of the geranylgeranylation of K-RasB occurred only at high concentrations of BZA- 2B. All of these properties of K-RasB were traced to the combined effects of its COOH-terminal CVIM sequence and the adjacent polylysine sequence, neither of which is present in H-Ras. These studies provide a potential explanation for the resistance of nontransformed cells to growth inhibition by BZA-5B. Inasmuch as the majority of Ras-related human cancers contain oncogenic versions of K-RasB rather than H-Ras, the current data suggest that in vitro studies of FTase inhibitors with potential anti-cancer activity should use authentic K-RasB as a substrate.",
author = "James, {G. L.} and Goldstein, {J. L.} and Brown, {M. S.}",
year = "1995",
doi = "10.1074/jbc.270.11.6221",
language = "English (US)",
volume = "270",
pages = "6221--6226",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
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T1 - Polylysine and CVIM sequences of K-RasB dictate specificity of prenylation and confer resistance to benzodiazepine peptidomimetic in vitro

AU - James, G. L.

AU - Goldstein, J. L.

AU - Brown, M. S.

PY - 1995

Y1 - 1995

N2 - BZA-5B, a benzodiazepine peptidomimetic, inhibits CAAX farnesyltransferase (FTase) and blocks attachment of farnesyl groups to oncogenic and wild-type H-Ras in animal cells. This compound slows the growth of cells transformed with oncogenic H-Ras at concentrations that do not affect the growth of nontransformed cells. This finding suggested that nontransformed cells may produce a form of Ras whose prenylation is resistant to BZA-5B. In the current studies, we found that FTase had a 50-fold higher affinity for K- RasB than for H-Ras in vitro. Farnesylation of K-RasB was inhibited by BZA- 2B, the active form of BZA-5B, but only at concentrations that were 8-fold higher than those that inhibited farnesylation of H-Ras. K-RasB, but not H- Ras, was also a substrate for CAAX geranylgeranyltransferase-1 (GGTase-1), and its affinity for the enzyme was equal to that of Rap1B, an authentic leucine-terminated substrate for GGTase-1. Inhibition of the geranylgeranylation of K-RasB occurred only at high concentrations of BZA- 2B. All of these properties of K-RasB were traced to the combined effects of its COOH-terminal CVIM sequence and the adjacent polylysine sequence, neither of which is present in H-Ras. These studies provide a potential explanation for the resistance of nontransformed cells to growth inhibition by BZA-5B. Inasmuch as the majority of Ras-related human cancers contain oncogenic versions of K-RasB rather than H-Ras, the current data suggest that in vitro studies of FTase inhibitors with potential anti-cancer activity should use authentic K-RasB as a substrate.

AB - BZA-5B, a benzodiazepine peptidomimetic, inhibits CAAX farnesyltransferase (FTase) and blocks attachment of farnesyl groups to oncogenic and wild-type H-Ras in animal cells. This compound slows the growth of cells transformed with oncogenic H-Ras at concentrations that do not affect the growth of nontransformed cells. This finding suggested that nontransformed cells may produce a form of Ras whose prenylation is resistant to BZA-5B. In the current studies, we found that FTase had a 50-fold higher affinity for K- RasB than for H-Ras in vitro. Farnesylation of K-RasB was inhibited by BZA- 2B, the active form of BZA-5B, but only at concentrations that were 8-fold higher than those that inhibited farnesylation of H-Ras. K-RasB, but not H- Ras, was also a substrate for CAAX geranylgeranyltransferase-1 (GGTase-1), and its affinity for the enzyme was equal to that of Rap1B, an authentic leucine-terminated substrate for GGTase-1. Inhibition of the geranylgeranylation of K-RasB occurred only at high concentrations of BZA- 2B. All of these properties of K-RasB were traced to the combined effects of its COOH-terminal CVIM sequence and the adjacent polylysine sequence, neither of which is present in H-Ras. These studies provide a potential explanation for the resistance of nontransformed cells to growth inhibition by BZA-5B. Inasmuch as the majority of Ras-related human cancers contain oncogenic versions of K-RasB rather than H-Ras, the current data suggest that in vitro studies of FTase inhibitors with potential anti-cancer activity should use authentic K-RasB as a substrate.

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