Expression and purification of beefy meaty peptide in Pichia pastoris

Abstract

We established a gene expression process to produce a flavor peptide in Pichia pastoris. The octapeptide Lys-Gly-Asp-Glu-Glu-Ser-Leu-Ala was isolated from beef digested by papain, known as "beefy meaty peptide" (BMP). It was considered a savory seasoning in previous literatures. To produce BMP by microorganisms, BMP expression structures encoding tandem repeats of the octapeptide were designed and cloned in E. coli DH5α, then integrated into the AOX1 gene of Pichia pastoris GS115. Clones containing different BMP-gene copies, with 4-copy, 8-copy, 12-copy and 16-copy, were obtained and expressed in Pichia pastoris GS115. For the ease of purification, 6×His tag was fused to the C-termini of the peptides. As a result, the fusion peptides were successfully purified by His-tag bind affinity resin. The BMP fusion peptides with expected sizes were secreted from the resulting strains of P. pastoris GS115.

Research progress in the screening and evaluation of umami peptides

“…(2020), and Wang et al. (2011) found umami taste after biological synthesis of the peptide. Zhang et al.…”

“…For example, Yamasaki-isolated beef meat peptides (BMPs) were umami (Yamasaki & Maekawa, 1978). Wassenaar et al (1995) found no umami taste after the chemical synthesis of the peptide, but Zeng et al (2010), Wu et al (2020), and Wang et al (2011) found umami taste after biological synthesis of the peptide. Zhang et al (2017) analyzed several possible reasons: first, the preparation method of umami peptides may affect the analysis of peptides; second, the isomers of peptides can also affect the taste of peptides; and, third, the difference in spatial structure interferes with the analysis of umami taste.…”

Safe preparation of beefy meaty peptide with Bacillus subtilis

“…The large‐scale production of bioactive peptides through engineering bacteria is an attractive strategy in the industry (Wang et al , ; Da et al , ), which could effectively facilitate the development and application of related products. Here, based on the above results of cultivation in shake flask, the scale ability of 8BMP was investigated by performing fermentation in a 5‐L fermenter.…”

“…The first two methods are not suitable for industrial production due to their low yields or high costs, whereas the last is an attractive approach to produce BMP (Li, ; Wu et al , ; Xia et al , ). Nevertheless, up to now, there are few reports about preparing BMP in microorganisms, except the biological expression of BMP in Pichia pastoris ( P. pastoris ) and in Escherichia coli ( E. coli ) (Zeng et al , ; Wang et al , ; Zhang et al , ). Furthermore, the industrial production of BMP was nearly limited by the relatively complicated fermentation of P. pastoris and the potential insecurity of E. coli (Hussein & Omaye, ; Vital et al , ).…”