T. Satyanarayana

Phytases: Microbial Sources, Production, Purification, and Potential Biotechnological Applications

ABSTRACT:  The review deals with phytase-producing microorganisms along with optimum conditions for its production. Various methods used for purifying phytases and their characteristics are discussed. Heterologous gene expression, cost-effective large-scale phytase production, and various biotechnological applications of the enzyme in animal feed and food industries are also discussed.

Statistical optimization of the medium components by response surface methodology to enhance phytase production by Pichia anomala

Enhanced phytase yield was attained with the yeast Pichia anomala using response surface methodology. Interactions were studied with three variables, viz. glucose, beef extract and inoculum density using Central Composite Design. Highest enzyme yields were obtained when the glucose and beef extract concentrations in the medium were 2 and 0.5%, respectively. The inoculum density was found to be an insignificant variable. The concentrations of the variables considered to be optimum by ‘one-variable-at-a-time’ method (glucose 4%, beef extract 1.25%) were actually higher than that actually required for maximum enzyme production. The application of response surface methodology resulted in an enhancement in phytase production and also minimized the cost of production. This is the first report on use of response surface methodology to improve phytase yield by any yeast.

α-Amylase production by thermophilic Bacillus coagulans in solid state fermentation

The production of extracellular, thermostable α-amylase by Bacillus coagulans B49 was studied in solid state fermentation (SSF). Significant levels of amylase were produced in wheat bran moistened with tap water [22 707 (U/g dry bacterial bran)]/salt solution (25 692 U). A marked improvement in enzyme production was achieved by optimization of SSF conditions (6700 U). Variations in the phase of α-amylase production and tires were observed depending on the culture vessels (Flask - 25 000 U at 72 h; reactor - 26 350 U at 48 h) and conditions (static - 24946 U at 72 h; shake flask - 25 263 at 48 h). Maximum enzyme production (26 350 U) was recorded in an aerated reactor. Catabolite repression was not evident in SSF even at high glucose concentrations. Cultivation in large trays yielded an enzyme titre of 17515 U at 96 h.

Phytase production by the yeast, Pichia anomala

Pichia anomala, isolated from dried flower buds of Woodfordia fruticosa, produced a high activity of an intracellular phytase, at 68 U per g dry biomass, when grown at 20 °C for 24 h in a medium containing glucose (40 g l−1) and beef extract (10 g l−1) supplemented with Fe2+ (0.15 mM). Partially purified phytase was optimally active at 60 °C and pH 4 with a half life of 7 days at 60 °C. It retained 85% of its activity at 80 °C for 15 min. The enzyme is suitable for supplementing animal feeds to improve the availability of phosphate from phytate.

Microbial glucoamylases: characteristics and applications

Glucoamylase is one of the oldest and widely used biocatalysts in food industry. The major application of glucoamylase is the saccharification of partially processed starch/dextrin to glucose, which is an essential substrate for numerous fermentation processes and a range of food and beverage industries. Glucoamylase for commercial purposes has traditionally been produced employing filamentous fungi, although a diverse group of microorganisms is reported to produce glucoamylase, since they secrete large quantities of the enzyme extracellularly. The commercially used fungal glucoamylases have certain limitations such as moderate thermostability, acidic pH requirement, and slow catalytic activity that increase the process cost. Consequently, the search for newer glucoamylases and protein engineering to improve pH and temperature optima leading to amelioration in catalytic efficiency of existing enzymes have been the major areas of research over the years. The present review focuses attention on the recent advances in molecular biology and protein engineering of glucoamylase to improve its production and functional properties including the so far success achieved in isolating mutants with enhanced thermostability and selectivity, higher pH optimum and improved catalytic activity. A comprehensive account is included on the diversity, regulation of production, classification, purification and properties, and potential applications of microbial glucoamylases to provide an overview on all the important aspects of the enzyme.

Cloning, expression and applicability of thermo-alkali-stable xylanase of Geobacillus thermoleovorans in generating xylooligosaccharides from agro-residues.

A xylanase gene (xyl-gt) of 1.224 kbp was cloned from the extremely thermophilic bacterium Geobacillus thermoleovorans that encodes a protein containing 408 amino acid residues. Eight conserved regions (signature sequences) of GH family 10 xylanases have been found in the xylanase. When the xylanase gene was cloned and expressed in Escherichia coli BL21 (DE3), the recombinant strain produced xylanase titer of 270 U mg(-1) which is 27-fold higher than the wild strain. It is optimally active at 80°C and pH 8.5 with a high thermostability over broad range of pH (6-12) and temperature (40-100°C). The end products of the hydrolysis of birch wood xylan and agro-residues included xylobiose, xylotriose, xylotetraose and xylopentaose. The xylanase of G. thermoleovorans is one of the rare xylanases that exhibits thermo-alkali-stability, and thus, it is a suitable candidate for pre-bleaching of paper pulps and generating xylooligosaccharides from agro-residues for use as prebiotics.

Statistical optimization of a high maltose‐forming, hyperthermostable and Ca2+‐independent α‐amylase production by an extreme thermophile Geobacillus thermoleovorans using response surface methodology

Aim: Statistical optimization for maximum production of a hyperthermostable, Ca2+‐independent and high maltose‐forming α‐amylase by Geobacillus thermoleovorans.

A marked enhancement in phytase production by a thermophilic mould Sporotrichum thermophile using statistical designs in a cost‐effective cane molasses medium

Aims:  Statistical optimization of phytase production by a thermophilic mould Sporotrichum thermophile in a cost‐effective cane molasses medium.

Microbial phytases in phosphorus acquisition and plant growth promotion

Phosphorus (P) is one of the major constituents in energy metabolism and biosynthesis of nucleic acids and cell membranes with an important role in regulation of a number of enzymes. Soil phosphorous is an important macronutrient for plant growth. Phosphorus deficiency in soil is a major problem for agricultural production. Total soil P occurs in either organic or in organic form. Phytic acid as phytate (salts of phytic acid) is the major form of organic phosphorus in soil and it is not readily available to plants as a source of phosphorus because it either forms a complex with cations or adsorbs to various soil components. Phosphate solubilizing microorganisms are ubiquitous in soils and could play an important role in supplying P to plants. Microorganisms utilizing phytate are found in cultivated soils as well as in wetland, grassland and forest soils. Various fungi and bacteria (including plant growth promoting rhizobacteria) hydrolyze this organic form of phosphorus secreting phosphatases such as phytases and acidic/alkaline phosphatases. A large number of transgenic plants have been developed which were able to utilize sodium phytate as sole source of phosphorus. However, the recombinant phytases were similar to their wild type counterparts in terms of their properties. Increased phytase/phosphatase activity in transgenic plants may be an effective approach to promote their phytate-phosphorus utilization. The extracellular phytase activity of transgenic plant roots is a significant factor in the utilization of phosphorus from phytate. Furthermore, this indicated that an opportunity exists for using gene technology to improve the ability of plants to utilize accumulated forms of soil organic phosphorus. This review is focused on the role of phytases and phytase producing microbes in promoting the growth of different plants.

A cost-effective cane molasses medium for enhanced cell-bound phytase production by Pichia anomala.

Aim:  Formulation of an inexpensive cane molasses medium for improved cell‐bound phytase production by Pichia anomala.