Isolation and characterization of the thylakoid membranes from the NaCl-resistant (NaCl(r)) mutant strain of the cyanobacterium Anabaena variabilis.

Abstract

NaCl-induced changes in the thylakoid membrane of wild-type Anabaena variabilis and its NaCl(r) mutant strain have been studied. Biochemical characterization of the thylakoid membrane was done by taking its absorption and fluorescence spectra at different wavelength. The thylakoid membranes of both strains were isolated by mechanical disruption of the freeze-dried and lysozyme-treated cells, followed by differential and density gradient centrifugation. The light absorption spectra of the thylakoid membrane showed three and two peaks in NaCl(r) mutant strain and its wild-type counterpart respectively at wavelengths of 400-850 nm. These peaks revealed that the thylakoid membrane contains a large amount of carotenoid and chlorophyll a. Fluorescence emission spectra of thylakoid membrane of NaCl(r) mutant and its wild-type strain at excitation wavelength of 335 nm showed two different peaks, one at 340 nm and the other at 663 nm respectively. The light absorption and fluorescence spectra of the thylakoid membrane also revealed that the membrane contained carotenoid pigment, chlorophyll (Chl) a, and a pigment with an emission peak at 335 nm. The HPLC analysis of the pigments of the thylakoid membrane indicates that the NaCl(r) mutant strain under NaCl stress contained an additional peak for the carotenoid pigment, which was lacking in its wild-type counterpart. The major peak in thylakoid membrane was that of echinenone and beta-carotene. Whereas the polypeptide composition of thylakoid membrane differed in the wild-type and its NaCl(r) mutant strain, no difference in the cell wall protein pattern was observed in both strains. The thylakoid membrane of NaCl(r) mutant strain contained two additional protein bands that were absent in its wild-type counterpart. The thylakoid membrane of the wild-type and its NaCl(r) mutant strain also showed morphological variations under NaCl stress.