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Biomedical subjects

Joseph Chappell

Publications and source records attributed to Joseph Chappell.

4 recordsLinked to original sources
PubMed2005

Surrogate splicing for functional analysis of sesquiterpene synthase genes.

A method for the recovery of full-length cDNAs from predicted terpene synthase genes containing introns is described. The approach utilizes Agrobacterium-mediated transient expression coupled with a reverse transcription-polydeoxyribonucleotide chain reaction assay to facilitate expression cloning of processed transcripts. Subsequent expression of intronless cDNAs in a suitable prokaryotic host provides for direct functional testing of the encoded gene product. The method was optimized by examining the expression of an intron-containing beta-glucuronidase gene agroinfiltrated into petunia (Petunia hybrida) leaves, and its utility was demonstrated by defining the function of two previously uncharacterized terpene synthases. A tobacco (Nicotiana tabacum) terpene synthase-like gene containing six predicted introns was characterized as having 5-epi-aristolochene synthase activity, while an Arabidopsis (Arabidopsis thaliana) gene previously annotated as a terpene synthase was shown to possess a novel sesquiterpene synthase activity for alpha-barbatene, thujopsene, and beta-chamigrene biosynthesis.

Alternative Splicing↗
PubMed2004

Characterization of botryococcene synthase enzyme activity, a squalene synthase-like activity from the green microalga Botryococcus braunii, Race B.

The extracellular matrix of the alga Botryococcus braunii, Race B, consists mainly of botryococcenes, which have potential as a hydrocarbon fuel. Botryococcenes are structurally similar to squalene raising the possibility of a common enzyme for the biosynthesis of both. While B. braunii squalene synthase (SS) enzyme activity has been documented, botryococcene synthase (BS) enzyme activity has not been. In the current study, an assay for BS activity has been developed and used to show that many of the assay conditions for BS enzyme activity are similar to those of SS. However, SS enzyme activity is stimulated by Tween 80 while BS enzyme activity is inhibited. Moreover, BS enzyme activity was correlated with the accumulation of botryococcenes during a B. braunii culture growth cycle, which was distinctly different from the profile of SS enzyme activity. While the current results indicate a conservation of enzymological features amongst the BS and SS enzymes, raising the possibility of one enzyme capable of catalyzing both activities, they are also consistent with these two activities arising from separate and distinct enzymes.

Chlorophyta↗
PubMed2002

Overexpression in Catharanthus roseus hairy roots of a truncated hamster 3-hydroxy-3-methylglutaryl-CoA reductase gene.

Catharanthus roseus (L.) G. Don hairy roots harboring hamster 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) (EC 1.1.1.88) cDNA without membrane-binding domain were evaluated by quantifying the levels of sterols and some indol-alkaloids. Clone 236, with the highest hybridization signal, had the lowest soluble and microsomal HMGR activity and produced more ajmalicine and catharanthine than the control but had reduced campesterol concentration. Clone 19, with low hybridization signal, had high soluble HMGR activity and produced high levels of campesterol and five to seven times more serpentine than the control but a low level of ajmalicine and no accumulation of catharanthine. These results suggest a possible role for HMGR in indole alkaloid biosynthesis and a possible cosuppression of both the endogenous and foreign HMGR genes in clone 236.

Animals↗