Comparison of the effects of trypsin and chymotrypsin on thylakoid membrane function

Raines, Christine Anne (1985) Comparison of the effects of trypsin and chymotrypsin on thylakoid membrane function. PhD thesis, University of Glasgow.

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The thylakoid membranes are the functional units upon which the photosynthetic energy-tansducing reactions occur. There is a strict relationship between the structure of these membranes and their functional capabilities; this is because the components of the photosynthetic electron-transport chain are bound to proteins in the membrane in a specific manner. Functional changes can be induced by using protediytic enzymes to alter the structure of the proteins. Two enzymes, trypsin (which digests bonds involving arginine and lysine) and chymotrypsin (which digest bonds involving tryptophan, tryosine and phenylalanine) have been used to investigate the functions of the thylakoid membrane. The rationale was that because these enzymes have different specificities for the bonds which they attack they may induce different functional changes in the membrane. The effects of these enzymes are compared. Measurements of light-induced oxygen evolution rates in the presence of exogenous electron acceptors showed that both trypsin and chymotrypsin altered the electron transport reactions in a number of ways. At higher concentrations both trypsin and chymotrypsin treatment of thylakoids caused the rate of electron transport to be dependent on the type of electron acceptor present and rendered electron transport insensitive to 3-(3,4-dichlorophenyl)-1,1-dimethylurea which inhibits photosystem II. These effects of trypsin have previously been shown to be due to partial digestion of a 32KDa protein on the acceptor side of photosystem II. Thus chymotrypsin may also affect this polypeptide. At lower concentrations trypsin, and to a lesser extent chymotrypsin, stimulated the rate of electron transport. This suggested that both of these enzymes tend to uncouple the thylakoid membranes. Trypsin and chymotrypsin altered the flash-induced field indicating absorption change measured at 520 nm (the electrochromic bandshift) in two ways : 1) the half-time of decay was decreased suggesting increased trans-membrane ion flux and 2) the extent was reduced. The decrease in the half-time of the decay of the electrochromic bandshift was correlated with uncoupling, by trypsin and to a lesser extent by chymotrypsin, of the thylakoid membrane. This proposal was supported by phosphorylation measurements which showed that both trypsin and chymotrypsin can, at least partially, inhibit ATP synthesis. But after brief trypsin incubation in the light a stimulation in the rate of phosphorylation and oxygen evolution (water to methyl viologen) was evident. It is possible that trypsin affects the e sub-unit, believed to be an inhibitor of ATPase function, however, no evidence to support this proposal was seen in polyacrylamide gel electrophoresis analysis of the ATPase. (Abstract shortened by ProQuest.).

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Molecular biology
Date of Award: 1985
Depositing User: Enlighten Team
Unique ID: glathesis:1985-76571
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 14:07
Last Modified: 19 Nov 2019 14:07

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