Euphorbia tirucalli β-Amyrin Synthase: Critical Roles of Steric Sizes at Val483 and Met729 and the CH-π Interaction between Val483 and Trp534 for Catalytic Action.

The functions of Val483, Trp534, and Met729 in Euphorbia tirucalli β-amyrin synthase were revealed by comparing the enzyme activities of site-directed mutants against that of the wild type. The Gly and Ala variants with a smaller bulk size at position 483 predominantly afforded monocyclic camelliol C, which suggested that the orientation of the (3S)-2,3-oxidosqualene substrate was not appropriately arranged in the reaction cavity as a result of the decreased bulk size, leading to failure of its normal folding into the chair-chair-chair-boat-boat conformation. The Ile variant, with a somewhat larger bulk, afforded β-amyrin as the dominant product.

Functional analysis of human aromatic amino acid transporter MCT10/TAT1 using the yeast Saccharomyces cerevisiae.

Tryptophan is an essential amino acid in humans and an important serotonin and melatonin precursor. Monocarboxylate transporter MCT10 is a member of the SLC16A family proteins that mediates low-affinity tryptophan transport across basolateral membranes of kidney, small intestine, and liver epithelial cells, although the precise transport mechanism remains unclear. Here we developed a simple functional assay to analyze tryptophan transport by human MCT10 using a deletion mutant for the high-affinity tryptophan permease Tat2 in Saccharomyces cerevisiae.

β-Amyrin synthase from Euphorbia tirucalli. Steric bulk, not the π-electrons of Phe, at position 474 has a key role in affording the correct folding of the substrate to complete the normal polycyclization cascade.

β-Amyrin, a triterpene, is widely distributed in plants and its glycosides confer important biological activities. Mutagenesis studies on β-amyrin synthase are very limited as compared with those of squalene-hopene cyclase and lanosterol synthase. This study was conducted to elucidate the function of the F474 residue of Euphorbia tirucalli β-amyrin cyclase, which is highly conserved in the superfamily of oxidosqualene cyclases.

Effect of cation-π interactions and steric bulk on the catalytic action of oxidosqualene cyclase: a case study of Phe728 of β-amyrin synthase from Euphorbia tirucalli L.

The function of the active-site residues of oxidosqualene cyclases (OSCs) has been presumed mainly in light of the product distribution; however, not much research has been performed into the enzymatic activity of mutated OSCs. β-Amyrin, which is widely found in the plant kingdom, is classified as an OSC; mutational studies on β-amyrin cyclase are very limited. Six site-specific mutations targeted at the Phe728 residue of Euphorbia tirucalli β-amyrin synthase (EtAS) were constructed to inspect the function of this aromatic residue.

Purification, kinetics, inhibitors and CD for recombinant β-amyrin synthase from Euphorbia tirucalli L and functional analysis of the DCTA motif, which is highly conserved among oxidosqualene cyclases.

β-Amyrin, a natural triterpene, is widely distributed in the plant kingdom, and its pentacyclic skeleton is produced by oxidosqualene cyclase (OSC). OSC enzymes are classified as membrane proteins, and they catalyze the polycyclization reaction of (3S)-2,3-oxidosqualene to yield nearly 150 different cyclic triterpene skeletons. To date, no report has described the successful purification and characterization of plant β-amyrin synthase.