Biology subjects

Zheng, F.

Publications and source records attributed to Zheng, F..

2 recordsLinked to original sources

bioRxiv2018

Characterizing Activity and Thermostability of GH5 Cellulase Chimeras from Mesophilic and Thermophilic Parents

Cellulases from glycoside hydrolase (GH) family 5 are key enzymes in the degradation of diverse polysaccharide substrates and are used in industrial enzyme cocktails to break down biomass. The GH5 family shares a canonical ({beta})8-barrel structure, where each ({beta}) module is essential for the enzyme stability and activity. Despite their shared topology, the thermostability of GH5 enzymes can vary significantly, and highly thermostable variants are often sought for industrial applications. Based on a previously characterized thermophilic GH5 cellulase from Talaromyces emersonii (TeEgl5A, with an optimal temperature of 90{degrees}C), we created ten hybrid enzymes with the mesophilic cellulase from Prosthecium opalus (PoCel5) to determine which elements are responsible for enhanced thermostability. Five of the expressed hybrid enzymes exhibit enzyme activity. Two of these hybrids exhibited pronounced increases in the temperature optima (10 and 20{degrees}C), T50 (15 and 19{degrees}C), Tm (16.5 and 22.9{degrees}C), and extended half life, t1/2 (~240- and 650-fold at 55{degrees}C) relative to the mesophilic parent enzyme, and demonstrated improved catalytic efficiency on selected substrates. The successful hybridization strategies were validated experimentally in another GH5 cellulase from Aspergillus nidulans (AnCel5), which demonstrated a similar increase in thermostability. Based on molecular dynamics simulations (MD) of both PoCel5 and TeEgl5A parent enzymes as well as their hybrids, we hypothesize that improved hydrophobic packing of the interface between 2 and 3 is the primary mechanism by which the hybrid enzymes increase their thermostability relative to the mesophilic parent PoCel5.\n\nIMPORTANCEThermal stability is an essential property of enzymes in many industrial biotechnological applications, as high temperatures improve bioreactor throughput. Many protein engineering approaches, such as rational design and directed evolution, have been employed to improve the thermal properties of mesophilic enzymes. Structure-based recombination has also been used to fuse TIM-barrel fragments and even fragments from unrelated folds, to generate new structures. However, there are not many research on GH5 cellulases. In this study, two GH5 cellulases, which showed TIM-barrel structure, PoCel5 and TeEgl5A with different thermal properties were hybridized to study the roles of different ({beta}) motifs. This work illustrates the role that structure guided recombination can play in helping to identify sequence function relationships within GH5 enzymes by supplementing natural diversity with synthetic diversity.

bioengineering↗

bioRxiv2018

Comparison of Generally Recognized as Safe Organic Acids for Disinfecting Fresh-cut Lettuce

In this study, we aimed to determine the organic acids (acetic, lactic, citric, malic, propionic, succinic, and tartaric acids; 1% and 0.5%, w/w or v/v) that were most effective for fresh-cut lettuce disinfection based on analysis of quality (i.e., color, electrolyte leakage, and sensory quality) and microbial examination. The results showed that these acids did not negatively affect the color quality (i.e., L*, a*, b*, whiteness index, and sensory color). Additionally, 0.5% lactic acid led to the lowest electrolyte leakage (0.83%), which was not significantly different (p > 0.05) from that of distilled water (0.46%). Lactic acid (1%) did not affect the sensory quality and led to the highest microbial reduction (1.45 log reduction in aerobic plate counts [APCs]; 2.31 log reduction in molds and yeasts [M&Y]) and was therefore recommended as the primary choice for lettuce disinfection. Malic acid (0.5%), with a 1.07% electrolyte leakage rate, 0.73 log reduction in APCs, and 1.40 log reduction in M&Y, was better than the other six acids (0.5%) and was recommended as a pH regulator and a potential synergistic agent for oxidizing sanitizers. Acetic acid (1%) negatively affected the sensory quality and led to the highest electrolyte leakage (2.90%). Microbial analysis showed that propionic acid (0.5% and 1%) was ineffective for disinfection of lettuce (p > 0.05); thus, acetic and propionic acids were not recommended. Our results provide insights into the choice of sanitizers and formula design in food safety.\n\nIMPORTANCESince chlorine is forbidden in several countries, generally recognized as safe organic acids are used in minimal processing industries and in household sanitizers. The disinfection efficacy of organic acids has been studied when used alone or with oxidizing sanitizers. However, since different antibacterial mechanisms, contact time, fresh produce, and concentration have been reported, the acids most effective for single fresh produce disinfection, especially that of lettuce, an important salad vegetable, are not known. Moreover, in developing countries, because of imperfections in field management, cold chain transportation, and minimal processing industry development, the demand for low-cost household sanitizers is greater than that for minimally processed fresh produce. In this work, microbial load in lettuce was determined after disinfecting with seven GRAS organic acids. The changes in quality were also determined. These results provide insights into the choice of minimal processing sanitizers and a formula design for household sanitizers.

microbiology↗