Paecilomyces variotii xylanase production, purification and characterization with antioxidant xylo-oligosaccharides production

Paecilomyces variotii xylanase was, produced in stirred tank bioreactor with yield of 760 U/mL and purified using 70% ammonium sulfate precipitation and ultra-filtration causing 3.29-fold purification with 34.47% activity recovery. The enzyme purity was analyzed on sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) confirming its monomeric nature as single band at 32 KDa. Zymography showed xylan hydrolysis activity at the same band. The purified enzyme had optimum activity at 60 °C and pH 5.0. The pH stability range was 5–9 and the temperature stability was up 70 °C. Fe2+and Fe3+ exhibited inhibition of xylanase enzyme while Cu2+, Ca2+, Mg2+ and Mn2+ stimulated its activity. Mercaptoethanol stimulated its activity; however, Na2-EDTA and SDS inhibited its activity. The purified xylanase could hydrolyze beechwood xylan but not carboxymethyl cellulose (CMC), avicel or soluble starch. Paecilomyces variotii xylanase Km and Vmax for beechwood were determined to be 3.33 mg/mL and 5555 U/mg, respectively. The produced xylanase enzyme applied on beech xylan resulted in different types of XOS. The antioxidant activity of xylo-oligosaccharides increased from 15.22 to 70.57% when the extract concentration was increased from 0.1 to 1.5 mg/mL. The enzyme characteristics and kinetic parameters indicated its high efficiency in the hydrolysis of xylan and its potential effectiveness in lignocellulosic hydrolysis and other industrial application. It also suggests the potential of xylanase enzyme for production of XOS from biomass which are useful in food and pharmaceutical industries.

Gene Editing of the Decoy Receptor LeEIX1 Increases Host Receptivity to Trichoderma Bio-Control

Fungal and bacterial pathogens generate devastating diseases and cause significant tomato crop losses worldwide. Due to chemical pesticides harming the environment and human health, alternative disease control strategies, including microorganismal bio-control agents (BCAs), are increasingly sought-after in agriculture. Bio-control microorganisms such as Trichoderma spp. have been shown to activate induced systemic resistance (ISR) in the host. However, examples of highly active bio-control microorganisms in agricultural settings are still lacking, due primarily to inconsistency in bio-control efficacy, often leading to widespread disease prior to the required ISR induction in the host. As part of its plant colonization strategy, Trichoderma spp. can secrete various compounds and molecules, which can effect host priming/ISR. One of these molecules synthesized and secreted from several species of Trichoderma is the family 11 xylanase enzyme known as ethylene inducing xylanase, EIX. EIX acts as an ISR elicitor in specific plant species and varieties. The response to EIX in tobacco and tomato cultivars is controlled by a single dominant locus, termed LeEIX, which contains two receptors, LeEIX1 and LeEIX2, both belonging to a class of leucine-rich repeat cell-surface glycoproteins. Both receptors are able to bind EIX, however, while LeEIX2 mediates plant defense responses, LeEIX1 acts as a decoy receptor and attenuates EIX induced immune signaling of the LeEIX2 receptor. By mutating LeEIX1 using CRISPR/Cas9, here, we report an enhancement of receptivity to T. harzianum mediated ISR and disease bio-control in tomato.