Structure and spectroscopy of methionyl-methionine for aquaculture.

The amino acid L-methionine is an essential amino acid and is commonly used as a feed supplement in terrestrial animals. It is less suitable for marine organisms because it is readily excreted. It is also highly water soluble and this results in loss of the feed and eutrophication of the water. To address these problems, the dipeptide DL-methionyl-DL-methionine (trade name: AQUAVI Met-Met) has been developed as a dedicated methionine source for aquaculture. The commercial product is a mixture of a racemic crystal form of D-methionyl-D-methionine/L-methionyl-L-methionine and a racemic crystal form of D-methionyl-L-methionine/L-methionyl-D-methionine. In this work, we have computationally, structurally, spectroscopically and by electron microscopy characterised these materials. The microscopy and spectroscopy demonstrate that there is no interaction between the DD-LL and DL-LD racemates on any length scale from the macroscopic to the nanoscale.

2-keto-4-(methylthio)butyric acid (keto analog of methionine) is a safe and efficacious precursor of L-methionine in chicks.

Relative bioefficacy and toxicity of Met precursor compounds were investigated in young chicks. The effectiveness of DL-Met and 2-keto-4-(methylthio)butyric acid (Keto-Met) to serve as L-Met precursors was quantified using Met-deficient diets of differing composition. Efficacy was based on slope-ratio and standard-curve methodology. Using L-Met as a standard Met source added to a purified diet, DL-Met and Keto-Met were assigned relative bioefficacy values of 98.5 and 92.5%, respectively, based on weight gain. Relative bioefficacy values of 98.5 and 89.3% were assigned to DL-Met and Keto-Met, respectively, when chicks were fed a Met-deficient, corn-soybean meal-peanut meal diet. Thus, both DL-Met and Keto-Met are effective Met precursor compounds in chicks. Additionally, growth-depressing effects of L-Met, DL-Met, and Keto-Met were compared using a nutritionally adequate corn-soybean meal diet supplemented with 15 or 30 g/kg of each compound. Similar reductions in weight gain, food intake, and gain:food ratio were observed for each compound. Subjective spleen color scores, indicative of splenic hemosiderosis, increased linearly (P < 0.01) with increasing intakes of each compound, suggesting a similarity in overall toxicity among these compounds. Because conversion of Keto-Met to L-Met in vivo merely requires transamination, Keto-Met may prove to be a useful supplement not only in food animal production, but also as a component of enteral and parenteral formulas for humans suffering from renal insufficiency.

Stereoselective synthesis of cis-p-menth-8-ene-1,7-diol, cis-p-menthane-1,7-diol, and cis-p-menthane-1,7,8-triol.

The natural products cis-p-menthane-1,7-diol (cis-IV), cis-p-menth-8-ene-1,7-diol (cis-I) and cis-p-menthane-1,7,8-triol (cis-II) are obtained starting from the corresponding cis-cyanohydrins, cis-2 and cis-7, respectively, by chemical transformation of the cyano into the hydroxymethyl group. The key step of the synthesis is the very high cis-selectivity (> or = 96 %) of the MeHNL-catalyzed HCN addition to 4-alkylcyclohexanones. From 4-isopropylcyclohexanone (1) the cyanohydrin cis-2 and from 4-(1-methylvinyl)cyclohexanone (6) the cyanohydrin cis-7 result almost quantitatively. Regioselective hydroxylation of cis-I affords the triol cis-II. X-ray crystal structure determinations of the final products confirm their cis-configuration.

Structure determinants of substrate specificity of hydroxynitrile lyase from Manihot esculenta.

Tryptophan 128 of hydroxynitrile lyase of Manihot esculenta (MeHNL) covers a significant part of a hydrophobic channel that gives access to the active site of the enzyme. This residue was therefore substituted in the mutant MeHNL-W128A by alanine to study its importance for the substrate specificity of the enzyme. Wild-type MeHNL and MeHNL-W128A showed comparable activity on the natural substrate acetone cyanohydrin (53 and 40 U/mg, respectively). However, the specific activities of MeHNL-W128A for the unnatural substrates mandelonitrile and 4-hydroxymandelonitrile are increased 9-fold and approximately 450-fold, respectively, compared with the wild-type MeHNL. The crystal structure of the MeHNL-W128A substrate-free form at 2.1 A resolution indicates that the W128A substitution has significantly enlarged the active-site channel entrance, and thereby explains the observed changes in substrate specificity for bulky substrates. Surprisingly, the MeHNL-W128A--4-hydroxybenzaldehyde complex structure at 2.1 A resolution shows the presence of two hydroxybenzaldehyde molecules in a sandwich type arrangement in the active site with an additional hydrogen bridge to the reacting center.