Journal ArticleDOI
A facile cell free synthesis of isotopically labeled (E)-4-hydroxy-3-methylbut-2-enyl diphosphate, a precursor of the plant hormone zeatin
TLDR
It is demonstrated that chromoplasts from Capsicum annuum, whose enzyme activity was impaired by freeze-thawing, accumulate 1.5 times more than normal, and this observation built the basis for the development of a cell-free system allowing the synthesis of this intermediate with labels in various positions.Abstract:
(E)-4-Hydroxy-3-methylbut-2-enyl diphosphate (1) is a key intermediate of the deoxyxylulose phosphate pathway of isoprenoid biosynthesis and a precursor of the plant hormone zeatin. The availability of this intermediate with various labeling patterns is pivotal for its use in biosynthetic studies. The number of positions, however, that can be easily labeled by chemical synthesis is limited, and the synthesis by means of recombinant enzymes is laborious and time consuming. We demonstrated that chromoplasts from Capsicum annuum, whose enzyme activity was impaired by freeze-thawing, accumulate 1. This observation built the basis for the development of a cell-free system allowing the synthesis of this intermediate with labels in various positions. With 2C-methyl-D-erythritol 2,4-cyclodiphosphate (5) as substrate, yields were in the range of 50%.read more
Citations
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Journal ArticleDOI
Preparation of Isotope Labeled/Unlabeled Key Intermediates in 2‐Methyl‐D‐erythritol 4‐Phosphate Terpenoid Biosynthetic Pathway
Heng Li,Shaobo Dai,Wen‐Yun Gao +2 more
TL;DR: Progress in chemical and enzymatic preparation of the key intermediates in this pathway is reviewed with the emphasis on the synthesis of 1-deoxy-D-xylulose 5-phosphate and 2-methyl- D-erythritol 4- phosphate with isotope labels.
References
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Journal ArticleDOI
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding
TL;DR: This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr with little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose.
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Biochemistry and Molecular Biology of the Isoprenoid Biosynthetic Pathway in Plants
TL;DR: The diverse nature of isoprenoids found in plants is summarized, emphasizing the wide range of physiological functions these compounds serve and some of the recently developed experimental systems that have helped researchers uncover much of the biochemistry and molecular biology of isOPrenoids.
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Arabidopsis CYP735A1 and CYP735A2 Encode Cytokinin Hydroxylases That Catalyze the Biosynthesis of trans-Zeatin
TL;DR: Root-specific induction of CYP735A2 expression by CK suggests that the trans-hydroxylation is involved in the regulation of CK metabolism and signaling in roots.
Journal ArticleDOI
The non-mevalonate pathway of isoprenoids: genes, enzymes and intermediates.
TL;DR: Although the mevalonate pathway had been considered for a long time as the unique source of biosynthetic isoprenoids, an alternative pathway has recently been discovered and the genes specifying these enzymes have been cloned from bacteria, plants and protozoa, opening the way to the identification of several novel pathway intermediates.
Journal ArticleDOI
Biosynthesis of terpenes: Studies on 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase
Petra Adam,Stefan Hecht,Wolfgang Eisenreich,Johannes Kaiser,Tobias Gräwert,Duilio Arigoni,Adelbert Bacher,Felix Rohdich +7 more
TL;DR: It is demonstrated that cell extract of an Escherichia coli strain engineered for hyperexpression of the ispH (lytB) gene catalyzes the in vitro conversion of 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate into IPP and DMAPP.