Microbial proteases, examples

Native ovoflavoprotein (49 kDa, pf=5.1) has, as does ovomucoid, certain antinutritional effects, as it inhibits serine proteases (trypsin, chymotrypsin and also microbial proteases) and has antiviral activity. Ovomacroglobulin (ovostatin) is an inhibitor of serine, cysteine, thiol and metalloproteases and shows antimicrobial activity. Some antinutritional effects are also seen in the basic glycoprotein avidin in raw egg white (relative molecular weight of the monomer is 15.6 kDa). It contains four identical subunits (pf = 9.5), each of which binds one molecule of biotin to give an unavailable complex. However, the denatured avidin, for example in hard-boiled eggs, does not interact with biotin. The interaction of riboflavin with flavoprotein (32 kDa, pf = 4.0) has, on the contrary, a positive influence on vitamin stability. Cystatin acts as cysteine protease inhibitor, and shows antimicrobial, antitumor and immunomodulating activities. 

Microorganisms produce a number of antibacterial substances of low molecular weight such as penicillin and streptomycin. In the past it was believed that microorganisms produce antibacterial substances to suppress their competitors. However, in the late 1960 s screening of enzyme inhibitors started, and most of enzyme inhibitors of microbial origin did not show any antibacterial activity. For example, protease inhibitors from Streptomyces, such as leupeptin and antipain, are very specific to enzyme inhibition and show no antibacterial effect. Therefore, it is more likely that these antibiotics and enzyme inhibitors have absolutely no role in the producing strains and that they are only the products of remnant genes that have lost their relevance over the course of evolution. 

CPA seems to occur only in mammals, but it should be noted that there is a related Zn endopeptidase, ther-molysin (EC 3.4.24.4), in thermophilic bacterium Bacillus thermoproteolyticus. Although its amino acid sequence and three-dimensional structure are unrelated to CPA. the active site structure is similar, and the mechanism of action also seems to be similar.This is an example of convergent evolution just like the case of serine proteases mammalian chymotrypsin and microbial subtilisin. 

Inhibitor peptides low molecular mass oligopeptide-fatty acid compounds of microbial origin which irreversibly inactivate plant and animal proteases. The inhibition is stoichiometric, i.e. 1 molecule I.p. inhibits 1 molecule enzyme. Examples are Leupeptin [acetyl-(or propionyl-)L-Leu-L-Leu-arginal the L-leu-cine can also be replaced by L-isovaline or L-valine], from Streptomyces species, inhibits cathepsin B, papain, trypsin, plasmin and cathepsin D, the effectiveness of the inhibition decreasing in that order. Pepsta-tin (isovaleryl-L-Val-L-Val-P-hydroxy-Y-NH2- -CH3-heptanoyl-L-Ala-P-hydroxy-Y-NHj-e-heptanoic acid), from actinomycetes, inhibits pepsin and cathepsin D. Chymostatin inhibits all known chymotrypsin types, cathepsin A, B, and D and papain. Antipain inhibits papain trypsin and plasmin. 

Methods for eliminating bitter peptides in partial protein hydrolysates are known, but they cause a significant loss of essential amino acids. These procedures usually include additional enzymatic hydrolysis under controlled conditions (a shorter time for the hydrolysis leads to higher peptides that are not bitter) and a selection of suitable proteases, such as aminopeptidases, carboxypeptidases and some other proteases. Enzymes of plant and microbial origin have been successfully used for this purpose. For example, the intracellular peptidases from Lactococcus lactis ssp. cremoris and Brevibacterium linens, which have high proteolytic activity, successfully hydrolyse bitter peptides in cheeses. 

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