Production of subtilisin proteases in bacteria and yeast

Abstract

In this review, we discuss the progress in the study and modif ication of subtilisin proteases. Despite longstanding applications of microbial proteases and a large number of research papers, the search for new protease genes, the construction of producer strains, and the development of methods for their practical application are still relevant and important, judging by the number of citations of the research articles on proteases and their microbial producers. This enzyme class represents the largest share of the industrial production of proteins worldwide. This situation can explain the high level of interest in these enzymes and points to the high importance of designing domestic technologies for their manufacture. The review covers subtilisin classif ication, the history of their discovery, and subsequent research on the optimization of their properties. An overview of the classes of subtilisin proteases and related enzymes is provided too. There is a discussion about the problems with the search for (and selection of) subtilases from natural strains of various microorganisms, approaches to (and specif ics of) their modif ication, as well as the relevant genetic engineering techniques. Details are provided on the methods for expression optimization of industrial subtilases of various strains: the details of the most important parameters of cultivation, i. e., composition of the media, culture duration, and the inf luence of temperature and pH. Also presented are the results of the latest studies on cultivation techniques: submerged and solid-state fermentation. From the literature data reviewed, we can conclude that native enzymes (i. e., those obtained from natural sources) currently hardly have any practical applications because of the decisive advantages of the enzymes modif ied by genetic engineering and having better properties: e. g., thermal stability, general resistance to detergents and specif ic resistance to various oxidants, high activity in various temperature ranges, independence from metal ions, and stability in the absence of calcium. The vast majority of subtilisin proteases are expressed in producer strains belonging to different species of the genus Bacillus. Meanwhile, there is an effort to adapt the expression of these enzymes to other microbes, in particular species of the yeast Pichia pastoris.

Evaluating the adhesive potential of the newly isolated bacterial strains in research exploitation of plant microbial interaction

“…A further increase in temperature to 50°C resulted in the formation of considerable amount of biofilm by SH-19. This is a well-known phenomenon and occurs probably owing to the diversity of optimal temperatures of the isolates (Rozanov et al, 2021).…”

Versatility of subtilisin: A review on structure, characteristics, and applications

“…Due to a combination of characteristics such as thermostability, wide range of pH compatibility, and broad specificity, family S8 or more commonly known as subtilisin family has turned out to be one of the most well‐known classes of proteases. Subtilisin is generally isolated from a wide range bacterial sources of the Bacillus genus such as Bacillus subtilis, B. amyloliquefaciens, B. licheniformis, B. lentus, B. clausii , and B. alkalophilus 1 . Isolation of subtilisin‐like protease from microorganisms are also not limited to species such as Thermococcus kodakarensis and Thermoactinomyces vulgaris .…”

“…Subtilisin is generally isolated from a wide range bacterial sources of the Bacillus genus such as Bacillus subtilis, B. amyloliquefaciens, B. licheniformis, B. lentus, B. clausii, and B. alkalophilus. 1 Isolation of subtilisin-like protease from microorganisms are also not limited to species such as Thermococcus kodakarensis and Thermoactinomyces vulgaris. At first, it is suggested that all Ser peptidases would be homologous to chymotrypsin.…”

“…Such enzymes include proteolytic enzymes or protease that are found in both prokaryotic and eukaryotic organisms. Protease is an enzyme that degrade protein by hydrolyzing the bonds between the polypeptide chain 1 . Its mode of action by peptide bonds hydrolysis has grouped protease under the hydrolase class of enzymes.…”

“…Protease is an enzyme that degrade protein by hydrolyzing the bonds between the polypeptide chain. 1 Its mode of action by peptide bonds hydrolysis has grouped protease under the hydrolase class of enzymes. Sixty percent of the global market of hydrolytic enzymes for industrial enzymes application are accounted by proteases.…”