Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00790 (PGA)
 2,475 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Potential feed enzyme additives for ruminants were tested in vitro for their stability to ruminal microbial and gastrointestinal proteolysis. Four commercial preparations from Trichoderma longibrachiatum (A, B, C, and D) and one from an undisclosed source (E) were incubated up to 6 h with ruminal fluid taken from four lactating dairy cows before or 2 h after feeding. The stability of preparation B was also tested in the presence of pepsin at pH 3 and pancreatin at pH 7. Cellulase (EC 3.2.1.4), cellulose 1,4-beta-cellobiosidase (EC 3.2.1.91), beta-glucanase (EC 3.2.1.6), xylanase (EC 3.2.1.8), beta-glucosidase (EC 3.2.1.21), and beta-xylosidase (EC 3.2.1.37) activities were monitored throughout the incubations. Polysaccharidase activities of all enzyme preparations were remarkably stable in ruminal fluid taken after feeding. Ruminal fluid obtained before feeding inactivated the polysaccharidases in preparations B and D to a greater extent than ruminal fluid obtained after feeding. Cellulase and cellulose 1,4-beta-cellobiosidase activities were the least stable, declining (P < 0.05) by 35 and 60% for preparations B and D, respectively. Xylanase activity of preparation D decreased (P < 0.05) by up to 30% after 6 h of incubation, whereas beta-glucanase activity was not affected. The ability to degrade exogenous enzymes also differed among cows (P < 0.05). Pepsin and acid (pH 3.0) did not affect polysaccharidases in preparation B but decreased glycosidase activities by 10 to 15% (P < 0.05) after 1 h of incubation. Pancreatin, at the maximum concentration used, inactivated cellulase, cellulose 1,4-beta-cellobiosidase, and xylanase activities at a rate of 0.55, 1, and 0.45%/min, respectively. beta-Glucosidase and beta-xylosidase activities decreased by 1 and 0.75%/min, respectively. Partial proteolysis of cellulase, cellulose 1,4-beta-cellobiosidase, and xylanase by pancreatin produced a transient increase in activity. This twofold increase for cellulase and fourfold increase for cellulose 1,4-beta-cellobiosidase was directly proportional to pancreatin concentration. These results suggest that the enzyme feed additives tested were stable in the rumen of animals after feeding. Exogenous enzymes are likely to be more susceptible to the host gastrointestinal proteases in the abomasum and intestines than to ruminal proteases. However, exogenous polysaccharidases may survive for a considerable period of time in the small intestine and they probably maintain activity against target substrates in this environment.
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PMID:Resistance of feed enzymes to proteolytic inactivation by rumen microorganisms and gastrointestinal proteases. 1142 1

Pichia pastoris (Komagataella sp.) is broadly used for the production of secreted recombinant proteins. Due to the high rate of protein production, incorrectly folded proteins may accumulate in the endoplasmic reticulum (ER). To restore their proper folding, the cell triggers the unfolded protein response (UPR); however, if the proteins cannot be repaired, they are degraded, which impairs process productivity. Moreover, a non-producing/non-secreting subpopulation of cells might occur, which also decreases overall productivity. Therefore, an in depth understanding of intracellular protein fluxes and population heterogeneity is needed to improve productivity. Under industrially relevant cultivation conditions in bioreactors, we cultured P. pastoris strains producing three different recombinant proteins: penicillin G acylase from Escherichia coli (EcPGA), lipase B from Candida antarctica (CaLB) and xylanase A from Thermomyces lanuginosus (TlXynA). Extracellular and intracellular product concentrations were determined, along with flow cytometry-based single-cell measurements of cell viability and the up-regulation of UPR. The cell population was distributed into four clusters, two of which were viable cells with no UPR up-regulation, differing in cell size and complexity. The other two clusters were cells with impaired viability, and cells with up-regulated UPR. Over the time course of cultivation, the distribution of the population into these four clusters changed. After 30 h of production, 60% of the cells producing EcPGA, which accumulated in the cells (50-70% of the product), had up-regulated UPR, but only 13% of the cells had impaired viability. A higher proportion of cells with decreased viability was observed in strains producing CaLB (20%) and TlXynA (27%). The proportion of cells with up-regulated UPR in CaLB-producing (35%) and TlXynA-producing (30%) strains was lower in comparison to the EcPGA-producing strain, and a smaller proportion of CaLB and TlXynA (<10%) accumulated in the cells. These data provide an insight into the development of heterogeneity in a recombinant P. pastoris population during a biotechnological process. A deeper understanding of the relationship between protein production/secretion and the regulation of the UPR might be utilized in bioprocess control and optimization with respect to secretion and population heterogeneity.
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PMID:Single-Cell Approach to Monitor the Unfolded Protein Response During Biotechnological Processes With Pichia pastoris. 3087 40