Cyclodextrin glucanotransferase: fundamentals and biotechnological implications.

Cyclodextrin glucanotransferase (CGTase) is an extracellular enzyme of the GH13 α-amylase family that catalyzes a unique intramolecular reaction known as cyclization to transform α-1, 4-glucans and similar starches into cyclodextrins. They also catalyze intermolecular transglycosylation reactions namely coupling, disproportionation, and some hydrolyzing effects on starch. The monomeric structures of the CGTase exhibit five domains (A, B, C, D, and E domains) with different molecular weights and amino acid sequences depending on the source. Among bacteria, Bacillus genus covers approximately 90% of the CGTase producers, while other genera like Klebsiella, Paenibacillus, and Thermoanaerobacter also shown decent contributions in recent studies. CGTase production is highly supported by alkaliphilic bacteria under submerged fermentation rather than solid-state fermentation. The bacterial sources, biochemical properties, production conditions, and structure of CGTases are compiled in this review. Cyclodextrins have the unique property of making inclusion complexes with various compounds, hence widely used in the food, pharmaceutical, cosmetics, laundry, and chemical sectors. This review presents a comprehensive view of CGTase produced by Bacillus spp., and other bacterial genera like Klebsiella, Paenibacillus, and Microbacterium. It also gives insight of the properties and recent biotechnological applications of cyclodextrins. KEY POINTS: • Transglycosylation reactions catalyzed by CGTase and their structural properties. • Comparative data of CGTase production by various genera and Bacillus spp. • Structures, properties, and applications of different cyclodextrins.

Understanding the Fermentation Potentiality For Gibberellic Acid (GA3) Production Using Fungi.

Gibberellins represent an important group of potent phytohormones, growth-promoting, closely related diterpenoid acids biologically derived from tetracyclic diterpenoid hydrocarbon. Among these, gibberellic acid (GA3) has received the greatest attention. GA3 is a highly valued plant growth regulator which has various applications in agriculture. It is extensively used for beneficial effects including stem elongation, elimination of dormancy, sex expression, seed germination, flowering, and fruit senescence. Along with plants, many microbes are also producing GA3 as their secondary metabolite, and among these, fungi are reported to produce a higher amount of GA3. Fermentation technology based on submerged fermentation and solid-state fermentation for the production of GA3 has been used with its merits and demerits using Fusarium moniliforme fungus in the industry. Several mathematical models and optimization tools were also designed for enhancing the fermentative yield by researchers. The detailed analysis is essential to understand all the fermentation aspects, various unit parameters, process operation approaches, reduction in cost, and assessment of the possible uses of these models in the production of GA3 for higher yield. Recently, exclusive research is executed to lower down the production cost of GA3 approaching various strategies.

Statistical optimization of medium components for biosurfactant production by Pseudomonas guguanensis D30.

Biosurfactant production by Pseudomonas guguanensis D30 was reported using mineral oil in submerged condition. Twelve medium components were tested at two levels by Plackett-Burman design, among them, mineral oil, yeast extract, peptone, MgSO4, and CaCl2 found significant on the basis of emulsification index. These five significant components were further optimized through central composite design (CCD). The experimental design was successfully used for regression analysis and the significant model suggested the solution of 10% (v/v) mineral oil, 3.0 g/L (w/v) yeast extract and 0.2 g/L (w/v) peptone for 13.14 g/L predicted biosurfactant production. We kept the suggested concentrations of medium components and got 13.34 ± 0.08 g/L biosurfactant production, which is almost double the conventional one-factor-at-a-time production (7.126 ± 0.12 g/L). It reduced the surface tension of the medium up to 28 ± 1.2 mN/m. We found ethyl acetate a suitable solvent for biosurfactant extraction amongst methanol, chloroform, and methanol:chloroform. The partially purified biosurfactant was chemically characterized as lipopeptide by Fourier transform infrared spectroscopy (FT-IR).

Plant growth promoting activities and effect of fermented panchagavya isolate Klebsiella sp. PG-64 on Vigna radiata.

A natural bacterial isolate from fermented panchagavya named as PG-64, exhibits multiple plant growth-promoting traits. This Gram-negative bacteria was identified as Klebsiella sp. PG-64 by 16S rRNA gene sequencing. The Klebsiella sp. PG-64 has shown production of indole acetic acid (106.0 µg/ml), gibberellic acid (20.0 µg/ml), ammonia (7.12 µmol/ml), exopolysaccharide (2.04% w/v) and phosphate solubilization (106.0 µg/ml). It produced 437 µg/ml IAA with 0.75% (w/v) L-tryptophan supplementation and was increased to 575 µg/ml in a laboratory-scale fermenter. The PG-64 has shown tolerance to abiotic stress conditions like pH (5.0-12.0), temperature (28-46 °C), salt (0.5-10.0% w/v NaCl) and osmotic resistance (1-10% w/v PEG-6000). The PG-64 also produced 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase (0.3 ng α-ketobutyrate/mg protein/h) indicating its potential for drought tolerance. Owing to its diverse properties, the effect of Klebsiella sp. PG-64 on Vigna radiata (Mung bean) was examined. The seeds treated with PG-64 culture showed 92% germination with a good seedling vigour index (202). In the pot study, Vigna radiata growth showed 2.23, 1.55, 2.00, 1.65, 1.73, 1.88, 5.00, 5.00, 1.57 times increase in primary root length, dry root weight, root hair numbers, leaf width, leaf numbers, leaf area, fruits number, flower number and chlorophyll content, respectively after 75 days. The application of Klebsiella sp. PG-64 culture resulted in substantial growth enhancement of Vigna radiata. The Klebsiella sp. PG-64 has multiple plant growth-promoting properties along with capabilities to tolerate abiotic stresses, making it a promising liquid biofertilizer contender for various crops.

Bio-prospecting the future in perspective of amidohydrolase L-glutaminase from marine habitats.

In the current scenario, considerable attention is being given to the enzyme L-glutaminase (EC 3.5.1.2). It belongs to the amidohydrolase class adherent to the family of serine-reliant ß-lactamases and the penicillin-binding proteins due to its higher affinity to polymerize and modify peptidoglycan synthesis. However, based on the catalytic proficiency, L-glutaminase is characterized as a proteolytic endopeptidase that cleaves peptide linkage and emancipates various byproducts, viz. ammonia along with glutamate. L-glutamine is considered the key amino acid reportedly involved in multiple metabolic pathways such as nitrogen metabolism. The present review is focused on the recent development and aspects concomitant to the biotechnological applicability of L-glutaminase predominantly from the marine habitat. Additionally, a majority of L-glutaminases finds application in cancer therapy as therapeutic agents, especially for acute lymphocytic leukaemia. The in vitro studies have been effective against various human cancer cell lines. L-glutaminase enhances the growth of probiotic bacteria. Apart from all these applications, it is suitably applicable in fermented foods as a flavour enhancer especially the umami flavour and content. Marine habitats have largely been exploited for their bio-catalytic potential but very scarcely for therapeutic enzymes. Some of the reports of such marine bacterial isolates from Bacillus sp., Pseudomonas sp. and Vibrio sp. are in the domain, but none highlights the therapeutic applications predominantly as anticancer and anti-proliferative agents. KEY POINTS: The exploration of marine habitats along the Gujarat coasts mainly for bacteria secreting L-glutaminase is scarcely reported, and even more scarce are the amidohydrolases from these marine niches as compared to their terrestrial counterparts. Microbial sourced amidohydrolase has wide bio-applicability that includes food, cosmetics and therapeutics especially as anticancer/anti-proliferative agent making it of immense biotechnological significance.

Valorization of agro-food wastes: Ease of concomitant-enzymes production with application in food and biofuel industries.

The novel and greener approach toward the co-production of hydrolytic enzymes in a single-cultivation medium with inexpensive substrates can bring down the production costs. Likewise, the natural and industrial organic biomass/solid are all nutritionally rich substrates waiting for free use in industries such as food, biofuel, etc. Valorization must broaden its applications in industries and households with a step towards a sustainable environment. The biofuel approach can be projected as one of the most promising deputations to meet future energy demands, in reduction of the environmental pollution due to excessive fossil fuel consumption. The present review highlights the multifaceted stature of microbial enzymes in this direction and possible implications mainly in the food industry and biofuel with the global impact of similar bio-based industries. In this review, design scale-up, fermentation cost, energy needs,and agro-food waste management have been meticulously delineated.

Microbial surfactants: A journey from fundamentals to recent advances.

Microbial surfactants are amphiphilic surface-active substances aid to reduce surface and interfacial tensions by accumulating between two fluid phases. They can be generically classified as low or high molecular weight biosurfactants based on their molecular weight, whilst overall chemical makeup determines whether they are neutral or anionic molecules. They demonstrate a variety of fundamental characteristics, including the lowering of surface tension, emulsification, adsorption, micelle formation, etc. Microbial genera like Bacillus spp., Pseudomonas spp., Candida spp., and Pseudozyma spp. are studied extensively for their production. The type of biosurfactant produced is reliant on the substrate utilized and the pathway pursued by the generating microorganisms. Some advantages of biosurfactants over synthetic surfactants comprise biodegradability, low toxicity, bioavailability, specificity of action, structural diversity, and effectiveness in harsh environments. Biosurfactants are physiologically crucial molecules for producing microorganisms which help the cells to grasp substrates in adverse conditions and also have antimicrobial, anti-adhesive, and antioxidant properties. Biosurfactants are in high demand as a potential product in industries like petroleum, cosmetics, detergents, agriculture, medicine, and food due to their beneficial properties. Biosurfactants are the significant natural biodegradable substances employed to replace the chemical surfactants on a global scale in order to make a cleaner and more sustainable environment.

Understanding the Basis of Occurrence, Biosynthesis, and Implications of Thermostable Alkaline Proteases.

The group of hydrolytic enzymes synonymously known as proteases is predominantly most favored for the class of industrial enzymes. The present work focuses on the thermostable nature of these proteolytic enzymes that occur naturally among mesophilic and thermophilic microbes. The broad thermo-active feature (40-80 °C), ease of cultivation, maintenance, and bulk production are the key features associated with these enzymes. Detailing of contemporary production technologies, and controllable operational parameters including the purification strategies, are the key features that justify their industrial dominance as biocatalysts. In addition, the rigorous research inputs by protein engineering and enzyme immobilization studies add up to the thermo-catalytic features and application capabilities of these enzymes. The work summarizes key features of microbial proteases that make them numero-uno for laundry, biomaterials, waste management, food and feed, tannery, and medical as well as pharmaceutical industries. The quest for novel and/or designed and engineered thermostable protease from unexplored sources is highly stimulating and will address the ever-increasing industrial demands.

Catalysis and stability of an alkaline protease from a haloalkaliphilic bacterium under non-aqueous conditions as a function of pH, salt and temperature.

A haloalkaliphilic bacterium, isolated from Coastal Gujarat (India) was identified as Oceanobacillus sp. (GQ162111) based on 16S rRNA gene sequence. The organism grew and secreted extra cellular protease in presence of various organic solvents. At 30% (v/v) concentration of hexane, heptane, isooctane, dodecane and decane, significant growth and protease production was evident. The alkaline protease was purified in a single step on phenyl sepharose 6 FF with 28% yield. The molecular mass as judged by SDS-PAGE was 30 kDa. The temperature optimum of protease was 50°C and the enzyme retained 70% activity in 10% (v/v) isooctane. Effect of salt and pH was investigated in combination to assess the effect of isooctane. In organic solvents, the enzyme was considerably active at pH 8-11, with optimum activity at pH 10. Salt at 2 M was optimum for activity and enzyme maintained significant stability up to 18 h even at 3 M salt concentration. Patters of growth, protease production, catalysis and stability of the enzyme are presented. The study resumes significance as limited information is available on the interaction of haloalkaliphilic bacteria and their enzymes with organic solvents.