Galacto-oligosaccharides (GOS) are prebiotics that have a beneficial effect on human health by promoting the growth of probiotic bacteria in the gut, in addition to having various applications in the food industry. GOS are generally produced from lactose in a reaction catalyzed by ß-galactosidase. Synthesis of GOS from whey permeate (WP) (ultrafiltration of whey, concentrated then spray dried) using surface engineered β-galactosidase in Pichia pastoris (P. pastoris) is a novel method to convert waste into a valuable product. Cell-surface display is the expression of peptides and proteins on the surface of living cells by fusing them to functional components of cells. Surface engineered cells have many potential uses. The Flo1p flocculation functional domain, thought to be located near the N terminus, recognizes and adheres non-covalently to cell-wall components such as α-mannan carbohydrates, causing reversible aggregation of cells into flocs.

Antimicrobial peptides (AMPs) are an important component of innate defense mechanisms with broad-spectrum activities against various pathogenic microorganisms, including Gram-positive and Gram-negative bacteria, fungi, and viruses. Antibiotic resistance has become a pervasive and global health burden, resulting in the immediate need to develop a new class of antibiotic substances. We screened a 16-mer random peptide library using the yeast two-hybrid system with Beclin 1 as bait and found that two 16-mer peptides (named P4 and P30) appeared to interact with Beclin1 in the β-gal assay. The two candidate cDNAs were introduced into the yeast secretory system of Pichia pastoris and their expression induced in the presence of methanol. Spectrophotometric analysis and Disc clear zone assay using the supernatant of the yeast growth media showed that both of the two peptides had strong activities against Staphylococcus aureus, MRSA (methicillin resistance Staphylococcus aureus), MRSA2242, and MRSA-2250, but no effect on commensal Lactobacillus strains. PCR analysis of the genomic DNA of transformed Pichia pastoris using AOX1 primers revealed that the two cDNAs were integrated into the genome at the AOX1 locus. Our result suggests that these peptides could be developed as a useful alternative to classic chemical antibiotics.