Structural and functional insights into the enzymatic activities of lipases from Burkholderia stagnalis and Burkholderia plantarii.

Lipases with high interesterification activity are important enzymes for industrial use. The lipase from Burkholderia stagnalis (BsL) exhibits higher interesterification activity than that from Burkholderia plantarii (BpL) despite their significant sequence similarity. In this study, we determined the crystal structure of BsL at 1.40 Å resolution. Utilizing structural insights, we have successfully augmented the interesterification activity of BpL by over twofold. This enhancement was achieved by substituting threonine with serine at position 289 through forming an expansive space in the substrate-binding site. Additionally, we discuss the activity mechanism based on the kinetic parameters. Our study sheds light on the structural determinants of the interesterification activity of lipase.

Analysing the substrate multispecificity of a proton-coupled oligopeptide transporter using a dipeptide library.

Peptide uptake systems that involve members of the proton-coupled oligopeptide transporter (POT) family are conserved across all organisms. POT proteins have characteristic substrate multispecificity, with which one transporter can recognize as many as 8,400 types of di/tripeptides and certain peptide-like drugs. Here we characterize the substrate multispecificity of Ptr2p, a major peptide transporter of Saccharomyces cerevisiae, using a dipeptide library. The affinities (Ki) of di/tripeptides toward Ptr2p show a wide distribution range from 48 mM to 0.020 mM. This substrate multispecificity indicates that POT family members have an important role in the preferential uptake of vital amino acids. In addition, we successfully establish high performance ligand affinity prediction models (97% accuracy) using our comprehensive dipeptide screening data in conjunction with simple property indices for describing ligand molecules. Our results provide an important clue to the development of highly absorbable peptides and their derivatives including peptide-like drugs.

Soy peptides enhance yeast cell growth at low temperatures.

PURPOSE OF WORK: To elucidate the mechanisms of high performance by soy peptides during yeast fermentation in low temperature stress conditions and to identify a type of soy peptide that is suitable for yeast fermentation at low temperatures during brewing. The growth of a tryptophan auxotrophic yeast strain in a medium containing soy peptide (SP) was compared with that in a medium containing an equivalent composition of free amino acid mixture. At low temperature (10 °C), the cells grew in the medium containing SP but not in the medium containing the free amino acid mixture. Free tyrosine and phenylalanine inhibited the uptake of free tryptophan at low temperatures whereas most of the tyrosine and phenylalanine were present as oligopeptides in SP. The cells could transport free tryptophan without being inhibited by free tyrosine and phenylalanine at low temperatures in the medium containing SPs. Soy peptide-AM that lacks free tyrosine and phenylalanine was more effective in promoting cell growth compared with a highly hydrolyzed version.

Soy peptides enhance heterologous membrane protein productivity during the exponential growth phase of Saccharomyces cerevisiae.

In this study, the production of eight G protein-coupled receptors by Saccharomyces cerevisiae was compared using two types of media, one of which contained soy peptides and the other free amino acids. Yeast cell growth improved in the medium with soy peptides, and the expression levels of six of the receptors increased during the exponential phase by an average of 2.3-fold as against the free amino acid-based medium. The enhancement of protein expression by soy peptides can be explained by alleviation of metabolite stress due to amino acid source depletion caused by heterologous protein expression.

Modification of yeast characteristics by soy peptides: cultivation with soy peptides represses the formation of lipid bodies.

We have previously reported that the cultivation of yeast cells with soy peptides can improve the tolerance of yeast to freeze-thaw stress (Izawa et al. Appl Microbiol Biotechnol 75:533-538, 2007), indicating that soy peptides can modify the characteristics of yeast cells. To gain a greater understanding of the potencies of soy peptides, we further investigated the effects of cultivation with soy peptides on yeast physiology and found that soy peptides repress the formation of lipid bodies (also called lipid droplets or lipid particles), in which neutral lipids are accumulated. Compared with casein peptone, bacto peptone, yeast nitrogen base, and free amino acid mixtures having the same amino acid composition as soy peptides, cultivation with soy peptides caused decreased levels of mRNAs of neutral lipid synthesis-related genes, such as DGA1, and repressed the formation of lipid bodies and accumulation of triacylglycerol. These results indicate that soy peptides affect the lipid metabolism in yeast cells, and also demonstrate a potentiality of edible natural ingredients as modifiers of the characteristics of food microorganisms.

Effect of soy peptide on brewing beer.

Here, we examined the effect of soy peptides (SPs) on the fermentation and growth of Yeast Bank Weihenstephan 34/70 (W34/70), a bottom-fermenting yeast. We compared fermentation for SP with that for a free amino acid (FAA) mixture having the same amino acid composition as SP, as a nitrogen source. Maltose syrup was used as a carbon source, and the medium contained excess amounts of essential minerals and vitamins. We observed that SP was better than FAA mixture at promoting fermentation and growth and that much more beta-phenylethyl alcohol was produced during fermentation with SP than with FAA mixture. Subsequently, we compared fermentations with the FAA mixture and selected mixtures containing various dipeptides of Phe as a nitrogen source. We found that the rates of Phe metabolism and beta-phenylethyl alcohol generation were much higher when Phe was presented as a dipeptide (Phe-Asp, Phe-Leu, or Phe-Phe) than when presented as FAA. These results show that amino acids such as Phe are absorbed more rapidly when presented as a peptide than as FAA, resulting in a more rapid production of beta-phenylethyl alcohol.