Microaerobic conditions enhance laccase production from Rheinheimera sp. in an economical medium.

Statistical optimization of aeration conditions viz. aerobic, microaerobic and anaerobic, was performed using response surface methodology (RSM) utilizing soybean meal as medium to enhance the production of laccase from Rheinheimera sp. Maximum laccase yield (18.48 × 105 U/L) was obtained under microaerobic (static) conditions sustained for 12 h in tandem with 26 h aerobically (150 rpm) grown culture, which was 17.03-fold higher than laccase production in the starting M162 medium under aerobic conditions (150 rpm). The reduction in incubation time from 72 to 38 h and utilization of cost-effective soybean meal as medium, which is easily available from local market, have provided a promising, eco-friendly method of laccase enzyme production. Enhanced expression of laccase gene under microaerobic conditions corresponded to the increased expression of fnr (fumarate nitrate reductase) gene, the oxygen sensing global regulator. The putative FNR-binding site upstream of laccase transcription initiation site was predicted to play an imperative role in Rheinheimera sp. adaptation from aerobic to microaerobic conditions and for enhanced laccase production.

Engineering multifunctional enzymes for agro-biomass utilization.

Lignocellulosic biomass is a plentiful renewable resource that can be converted into a wide range of high-value-added industrial products. However, the complexity of its structural integrity is one of the major constraints and requires combinations of different fibrolytic enzymes for the cost-effective, industrially and environmentally feasible transformation. An interesting approach is constructing multifunctional enzymes, either in a single polypeptide or by joining multiple domains with linkers and performing diverse reactions simultaneously, in a single host. The production of such chimera proteins multiplies the advantages of different enzymatic reactions in a single setup, in lesser time, at lower production cost and with desirable and improved catalytic activities. This review embodies the various domain-tailoring and extracellular secretion strategies, possible solutions to their challenges, and efforts to experimentally connect different catalytic activities in a single host, as well as their applications.

Antibiotic residues in environment: antimicrobial resistance development, ecological risks, and bioremediation.

The overuse of antibiotics and their disposal without processing are leading the environment and its inhabitants towards a serious health emergency. There is abundance of diverse antibiotic resistance genes and bacteria in environment, which demands immediate attention for the effective removal of antibiotics. There are physical and chemical methods for removal, but the generation of toxic byproducts has directed the efforts towards bioremediation for eco-friendly and sustainable elimination of antibiotics from the environment. Various effective and reliable bioremediation approaches have been used, but still antibiotic residues pose a major global threat. Recent developments in molecular and synthetic biology might offer better solution for engineering of microbe-metabolite biodevices and development of novel strains endowed with desirable properties. This review summarizes the impact of antibiotics on environment, mechanisms of resistance development, and different bioremediation approaches.

COVID-19 Diagnosis: Current and Future Techniques.

COVID-19 pandemic continues to be a global threat, affecting more than 200 countries/territories at both human and economic level. This necessitates the rapid development of highly reliable diagnostic methods in order to effectively and accurately diagnose the pathology to prevent the spread of COVID-19. Currently, RT-PCR is the most widely used method worldwide for SARS-CoV-2 detection. Serological assays are being used for sero-surveys of SARS-CoV-2 antibody prevalence in the community. Radiology imaging has been useful in the clinical diagnosis of COVID-19. These methods have their own limitations and there are continued efforts to develop easier, economic, highly sensitive and specific, point-of-care methods. Reverse transcription-loop mediated isothermal amplification (RT-LAMP), nucleic acid sequence-based amplification (NASBA), CRISPR-Cas-based detection, and digital PCR are such techniques being employed in research laboratories, with many awaiting diagnostic approval from competent authorities. This review highlights the rapidly expanding array of existing and in-development diagnostic tests/strategies that may be used to diagnose SARS-CoV-2 infection in both clinical and research settings.

Functional substitution of domain 3 (T1 copper center) of a novel laccase with Cu ions.

A bacterial laccase having potential to work in industry without mediator is of special interest. In this work, gene (1.83kb) encoding a novel laccase from Rheinheimera sp., having potential to deink waste paper without mediator, was cloned, over-expressed and the induced 69kDa protein (RhLacc) was purified and characterized. rhlacc gene was mutated by error prone PCR and mutants were sequenced. One mutant showed protein truncation resulting in the absence of domain 3 that contains T1 copper center. It is known that redox potential of T1 is the key parameter for substrate oxidation. Overexpression of this mutant gene showed induced 41.1kDa protein (∆RhLacc) that exhibited laccase activity but in the presence of added copper, compared to RhLacc which showed activity without added copper ions. Optimum temperature for both was 55°C. However, optimum pH varied with substrates. Kinetic studies showed ∆RhLacc had lower affinity for substrates except for guaiacol and reduced kcat in comparison to RhLacc. Both were able to deink old newspaper and degrade indigo carmine without mediator. The study suggests that the novel property to deink waste paper without mediator may not depend on the redox potential of T1 but other mechanisms using domains 1 and 2 may be involved.

Flavonoid-rich agro-industrial residues for enhanced bacterial laccase production by submerged and solid-state fermentation.

Laccases have potential applications in industrial, biotechnological, and environmental set ups. Development of cost effective and efficient production technologies has gained significant attention in recent years. To enhance the laccase production from Rheinheimera sp. (Gram negative) using submerged fermentation (SmF) and from Lysinibacillus sp. (Gram positive) using solid-state fermentation (SSF), the inducing effect of various flavonoid-rich agro-industrial residues was investigated. Peels of citrus fruits, soybean meal, tofu dreg, lignin monomers, and lingo-cellulosic waste, used tea leaves and peels of onion and kiwi, paper, and dying industry effluents were tested as inducers. In SmF, 0.1% of soybean meal, tofu dreg, and powdered orange peel were best, enhancing the laccase production 2.57-, 2.11-, and 2.05-fold, respectively. In SSF, 10 mg (w/w) of used tata acti green tea leaves per 5 g of wheat bran, 1% pulp and paper industry effluent (agro based), and 1% wine made from Sygium cumini enhanced the laccase production 2.69-, 2.61-, and 2.09-fold, respectively. These results suggest the utilization of these flavonoid and phenolic-rich waste materials to be potential enhancers of industrially important laccase production.