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  Double prenylation by RabGGTase can proceed without dissociation of the mono-prenylated intermediate

Thomä, N. H., Niculae, A., Goody, R. S., & Alexandrov, K. (2001). Double prenylation by RabGGTase can proceed without dissociation of the mono-prenylated intermediate. Journal of Biological Chemistry, 276(52): 1, pp. 48631-48636. Retrieved from http://www.jbc.org/cgi/content/abstract/276/52/48631.

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Genre: Journal Article
Alternative Title : J. Biol. Chem.

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 Creators:
Thomä, Nicolas H.1, Author
Niculae, Anca1, Author
Goody, Roger S.2, Author           
Alexandrov, Kirill2, Author           
Affiliations:
1Max Planck Institute of Molecular Physiology, Max Planck Society, ou_1753286              
2Abt. III: Physikalische Biochemie, Max Planck Institute of Molecular Physiology, Max Planck Society, ou_1753289              

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 Abstract: Rab geranylgeranyltransferase (RabGGTase) catalyzes the prenylation of Rab proteins. Despite possessing a single active site, RabGGTase is able to add geranylgeranyl moieties onto each of the two C-terminal cysteine residues of Rab. We have studied the kinetics of Rab double prenylation employing a combination of a novel high pressure liquid chromatography (HPLC)-based in vitro prenylation assay and fluorescence spectroscopy. Transfer of the first geranylgeranyl group proceeds with a k(1) = 0.16 s(-1), while the conversion from singly to double prenylated Rab is 4-fold slower (k(2) = 0.039 s(-1)). We found that following the first transfer reaction, the conjugated lipid is removed from the active site of RabGGTase but mono-prenylated Rab.REP complex remains bound to RabGGTase with a K-d < 1nM. In contrast to the doubly prenylated Rab7.REP dissociation of the mono-prenylated species from RabGGTase was only weakly stimulated by phosphoisoprenoid. Based on the obtained rate constants we calculated that at least 72% of mono-prenylated Rab molecules proceed to double prenylation without dissociating from RabGGTase. The obtained data provides an explanation of how RabGGTase discriminates between mono-prenylated intermediate and double prenylated reaction product. It also indicates that the phosphoisoprenoid acts both as a substrate and as a sensor governing the kinetics of protein-protein interactions in the double prenylation reaction.

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Language(s): eng - English
 Dates: 2001-12-28
 Publication Status: Published in print
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 Rev. Type: Peer
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Title: Journal of Biological Chemistry
  Alternative Title : J. Biol. Chem.
Source Genre: Journal
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Pages: - Volume / Issue: 276 (52) Sequence Number: 1 Start / End Page: 48631 - 48636 Identifier: -