Interrelated approach to optimization of biosynthesis and chemical isolation of biologically active substances: the production of penicillinamidase by Escherichia coli and peptidohydrolase by Xanthomonas sp.

The solution to the problem of optimizing conditions for the isolation of biologically active substances (BAS) from microbial cells should be based on investigations of the structural and functional characteristics of cultures. Models of two bacterial cultures, of Escherichia coli and a Xanthomonas species, producing enzymes the localization of which differ, is described. The isolation of membrane-bound penicillinamidase from E. coli was optimal in the 'preautolysis' period, when the components of the cytoplasm autolysed but the membranes remained intact. In contrast, the isolation of the cytoplasmic enzyme peptidohydrolase from Xanthomonas sp. was optimal during the period when the cell membranes markedly changed. Thus the physiological state of the cultures and the localization of the BAS within the cells are important determinants for optimization of the isolation process. It follows that all stages of a technological process for the production of BAS, i.e. biosynthesis, chemical isolation, etc., should be interrelated for a successful outcome.

Coagulation autolysis in microorganisms and its relation to coagulase production.

The phenomenon of coagulation autolysis was observed in two model microorganisms, i.e., a bacterial culture and an imperfect fungus. It was characterized by impairment of the cell membranes, followed by condensation and dehydration of the cytoplasm and long-term preservation of the cells in the form of coagulated cytoplasm. In this respect, it was similar to coagulation necrosis of human tissues. The autolysis in the microorganisms was accompanied by increase of their coagulase activity, the substrate specificity of the enzyme rather broad. The coagulase activity of the microorganisms was detected during the culture period between the lag-phase and the exponential growth phase, i.e., the phase of their active growth. It served as a signal to induce biosynthesis of peptidohydrolase and cleavage of proteins. We believe that the phenomenon of coagulation autolysis in these microorganisms is rather typical and can be considered as an adaptative reaction, inducing a cascade of events from synthesis of coagulase to overproduction of peptidohydrolases with proteolytic activity.

Acremonium chrysogenum differentiation and biosynthesis of cephalosporin.

On the basis of structure-functional analysis of the development of Acremonium chrysogenum, e.g. under conditions either stimulating antibiotic synthesis or not conductive to production, a scheme was proposed representing the various ways in which morphological differentiation occurs in the culture in dependence on the directions of its metabolism. Three types of culture differentiation were determined. Type 1 differentiation is characterized by the transition of the vegetative stage into the reproductive one with the formation of conidia. Type 2 differentiation is characterized by the formation of typical arthrospores also being the reproductive form. Type 3 differentiation is characterized by the multistage transformation of the mycelium organization into the yeast-like one which is metabolically more active and is a producer of antibiotics and enzymes. In addition to the defined regularities in the development and differentiation of Acremonium chrysogenum structural peculiarities were observed which could be helpful to the search for regulators or specific enzymes taking part in the culture development.

Influence of A-factor on the ultrastructure of the A-factor deficient mutant of Streptomyces griseus.

The fine structure of the A-factor deficient mutant strain of Streptomyces griseus grown in the absence or in the presence of this factor was studied by transmission electron microscopy. The submerged 96 h culture grown in the presence of the A-factor was characterized by the formation of a large amount of spores accompanied by mycelium disintegration. A number of essential ultrastructural changes were observed in the mycelium in the course of development and streptomycin biosynthesis after the A-factor addition. Thus, the cell wall was transformed in another manner at various developmental stages. For a long time the associated ribosomes were preserved. The number of intracytoplasmic membrane structures increased. New, unusual organelles appeared which consisted of parallel cylindrical structures. The possible nature of these organelles is discussed.