Novel Bacillus Milk-Clotting Enzyme Produces Diverse Functional Peptides in Semihard Cheese

Abstract Aspergillus oryzae RIB40 has 11 aspartic endopeptidase genes. We searched for milk-clotting enzymes based on the homology of the deduced amino acid sequence with chymosins. As a result, we identified a milk-clotting enzyme in A. oryzae. We expected other Aspergillus species to have a homologous enzyme with milk-clotting activity, and we found the most homologous aspartic endopeptidase from A. luchuensis had milk-clotting activity. Surprisingly, two enzymes were considered as vacuole enzymes according to a study on A. niger proteases. The two enzymes from A. oryzae and A. luchuensis cleaved a peptide between the 105Phe-106Met bond in κ-casein, similar to chymosin. Although both enzymes showed proteolytic activity using casein as a substrate, the optimum pH values for milk-clotting and proteolytic activities were different. Furthermore, the substrate specificities were highly restricted. Therefore, we expected that the Japanese traditional fermentation agent, koji, could be used as an enzyme source for cheese production.

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Screening of Edible Mushrooms Producing Milk-clotting Enzyme

Nippon Shokuhin Kagaku Kogaku Kaishi ◽

10.3136/nskkk.61.444 ◽

2014 ◽

Vol 61 (9) ◽

pp. 444-447 ◽

Cited By ~ 4

Author(s):

Kazuo Nakamura ◽

Nahoko Kobayashi ◽

Morimasa Tanimoto

Keyword(s):

Edible Mushrooms ◽

Milk Clotting ◽

Milk Clotting Enzyme

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Substrate specificity of the milk-clotting enzyme from Irpex lacteus on .KAPPA.- and .BETA.-casein.

Agricultural and Biological Chemistry ◽

10.1271/bbb1961.49.1621 ◽

1985 ◽

Vol 49 (6) ◽

pp. 1621-1631 ◽

Cited By ~ 6

Author(s):

Hideyuki KOBAYASHI ◽

Isao KUSAKABE ◽

Saeko YOKOYAMA ◽

Kazuo MURAKAMI

Keyword(s):

Substrate Specificity ◽

Irpex Lacteus ◽

Milk Clotting ◽

Milk Clotting Enzyme ◽

Beta Casein

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Influence of Amino Acid Substitutions in a Recombinant Cow Chymosin Molecule on the Dependence of its Coagulation Activity on the Concentration of Calcium Chloride in Milk

Biotekhnologiya ◽

10.21519/0234-2758-2021-37-1-81-86 ◽

2021 ◽

Vol 37 (1) ◽

pp. 81-86

Author(s):

A.V. Mironova ◽

V.A. Pushkarev ◽

J.G. Afanasieva ◽

D.N. Shcherbakov ◽

A.P. Rudometov ◽

...

Keyword(s):

Amino Acid ◽

Calcium Chloride ◽

Amino Acid Substitutions ◽

Salt Bridges ◽

Technological Properties ◽

Coagulation Activity ◽

Cheese Making ◽

Milk Clotting ◽

Milk Clotting Enzyme ◽

Milk Mixture

The influence of substitutions of amino acids involved in the formation of salt bridges on the cow chymosin technological properties has been studied. It was shown that point amino acid mutations Asp198→Lys, Asp156→Val and their combination decrease the sensitivity of coagulation activity of engineering recombinant chymosins to change in the concentration of calcium chloride in milk in the range of 1-5 mM. The found effect is a positive technological evidence in terms of cheese making, since it allows varying the content of calcium chloride added to the milk mixture without a threat of significant changes in the undesired proteolytic activity of used milk-clotting enzyme. recombinant chymosin, technological properties, amino acid substitutions, milk-clotting activity, concentration of calcium chloride. The work was performed within the framework of the state task of the Ministry of science and higher education of the Russian Federation (topic number fzmw-2020-0002, "Development of recombinant enzyme producers for cheese making").

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