Serine proteases, their inhibitors, and chitin synthetase in Phycomyces

Citation

Fischer, E. Peter (1977) Serine proteases, their inhibitors, and chitin synthetase in Phycomyces. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/4T38-3H24. https://resolver.caltech.edu/CaltechETD:etd-08042006-131928

Abstract

Phycomyces shows a positive growth response to blue light. Mutants are available which exhibit abnormal phototropism. They have been analysed genetically by complementation analysis. Seven complementation groups have been shown to be involved in the light information channel. They are designated mad A, -, G. Four groups are connected with the output end.

These findings encourage biochemical investigations. We make the hypothesis that light directly regulates the activity of chitin synthetase. It is investigated whether limited proteolysis represents one mode of such regulation.

Three serine proteases are isolated and characterized. The molecular weights are determined to be 18.000, 22.000, and 60.000 daltons. The proteases are solubilized by detergent or salt treatment. There are two specific soluble inhibitors present, which are also isolated and characterized. Both these proteins have a MW of 10.000 daltons. Each protease forms a 1:1 complex with its inhibitor. The inhibitors are present in excess in the cells. An acid protease is able to take the inhibitor off a serine protease-inhibitor complex. This protease has been partially purified.

The proteases and inhibitors of three mutant strains have been partially purified and compared with wild type. These mutants are disturbed at the output end of the light information channel. Mutant specific changes are detected but a connection to the changed behavioral responses has not been demonstrated.

Chitin synthetase activity is detected in three different fractions. One form appears to be soluble; the two particulate forms can be separated by density gradient centrifugation. The high density material is probably a plasma membrane fraction.

The serine proteases are able to activate all three forms of chitin synthetase.

A hypothetical cascade for chitin synthetase activation is discussed.

In an appendix the amino acid composition of cytochrome c of Phycomyces is presented.

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