Multifaceted in vivo and in vitro Characterization of cis-Prenyltransferase 5 from Solanum lycopersicum
Year: 2017
Type: dissertation
Abstract: The widespread occurrence of polyprenols throughout the plant kingdom is well documented. These compounds are believed to be assembled by a class of enzymes designated as cis-prenyltransferases (CPTs), which are encoded by small, yet largely uncharacterized, CPT gene families in plants. This research aimed to identify and characterize the CPT from Solanum lycopersicum (tomato) responsible for polyprenol synthesis. RNAi-mediated knockdown of one member of the tomato CPT family (SlCPT5) reduced polyprenols in leaves by ~70%. Assays with recombinant SlCPT5 determined that the enzyme synthesizes polyprenols of approximately C50-C55 in length and accommodates a variety of trans-prenyl diphosphate precursors as substrates. Introduction of SlCPT5 into the polyprenol-deficient yeast rer2∆ mutant resulted in the accumulation of C55 polyprenols in yeast cells, restored proper protein N-glycosylation, and rescued the temperature sensitive growth phenotype associated with its polyprenol deficiency. Finally, site-directed mutagenesis of SlCPT5 identified a suite of amino acids that are catalytically essential.
Author: Kevin A. Rea
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2757365591