Evaluation of the effect of Mutations on the Protease enzyme production by Lactobacillus delbrueckii

Author Name: Ankitha Mishra
Author Email: ankitamshr77@gmail.com

Abstract

Proteases are the enzymes produced in most organisms. Proteases produced by enzymatic methods are more environment friendly when compared to the chemical process and it has tremendous potential in the leather and several other industries. However optimization of protocol for the production of protease could involve several variables such as temperature, pH and incubation period. In the current work lactobacillus delbrueckii has been employed for the production of protease. Further the strain was subjected to mutations and the mutant strain was also employed in the production of the enzyme so as to check the effect of mutations on enzyme production. For the surprise it was found that the biochemical properties exhibited by the mutant strain of lactobacillus delbrueckii were exactly the same as shown by Neisseria flavescens. In other words the exposure of lactobacillus delbrueckii to UV Irradiated mutations made the bacteria to exhibit the properties of Neisseria flavescens. The current work includes the Isolation, Identification and Production of mutant strains of lactobacillus delbrueckii followed by the application of these strains in the production of the enzyme. The protocol also involves the quantitative estimation of the enzyme produced by the two strains.

Keywords

Spread plate technique, pure culture, Biochemical test, Lowry’s method, Salt precipitation, Dialysis, Enzyme kinetics.

Introduction

Enzymes are functional proteins, found within each of the cells, which help the cell to carry out its specialized functions. Proteases are enzymes that catalyze the breakdown of proteins. Protein breakdown is a normal process necessary to maintain cellular homeostasis. Active proteases can be found throughout your body, including the digestive tract, inside cells and circulating in the blood [1]. Proteases play a critical role in many physiological and pathological processes such as protein catabolism, blood coagulation, cell growth and migration, tissue arrangement, morphogenesis and development, inflammation, tumor growth and metastasis, activation of zymogens, release of hormones and pharmacologically active peptides from precursor proteins, and transport of secretory proteins across membranes [2].

Conclusion

The current work aimed to standardize and propose the method to improve the enzyme producing ability of lactobacillus delbrueckii. The wild strain of the bacteria was used to prepare the pure plates in two replicas. Once of the replica was directly used in the production of the enzyme whereas the other is subjected to mutations using UV irradiations.
In the next step both the mutant and wild strain were used for the production of the enzyme in vitro conditions. After extraction and purification of the enzymes produced by respective cultures the work was extended to compare the concentration of the enzyme produced by both the strains. The method employed for the enzyme estimation was Lowry’s method. The results showed that the enzyme produced from wild strain was 3 fold less than the mutant strain. The wild strain produced 0.23mg/ml of enzyme into the broth where as the mutant strain produced 0.70mg/ml.
These results conclude that the enzyme concentrating produced by the mutant strains was much higher than the wild strains. Thus induction of mutations would be a positive approach to enhance the yield of the enzyme by the cultures. Further the biochemical characterization of mutant strains showed that there exhibited the properties that were exactly identical to the properties of the bacteria Neisseria flavescens.
This data can be used to precede the research towards the comparison of the similarities exhibited by two strains lactobacillus delbrueckii and Neisseria flavescens.

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