| Abstract |
- The photosynthetic activities, the pigmentation and the chloroplast
membrane polypeptides of several mutants of the green alga, Scenedesmus
obliquus, with complete or limited deficiencies in the light-harvesting
complex of photosystem II are characterized and compared to those of the
normal phenotype.. The principal phenotype examined in this study, K0 -9,
behaves similarly to the WT Scenedesmus in that it grows autotrophically,
mixotrophically and heterotrophically; cells grown under these various
conditions retain near normal photosynthetic capacity without alteration of
either photosystem I or photosystem II activities. Also, the associated
chloroplast membrane polypeptides, as well as the polypeptides of the
apoproteins of the LHC-II, appear identical to those of the WT. However,
absorption spectra analyses and gel electrophoresis studies revealed that
the mutant lacks the major light-harvesting complex, LHCP-II ( as shown
in "green" gels) and the corresponding chlorophyll a and chlorophyll h of
the light-harvesting complex.
Examination of total pigment extracts of WT and K0 -9 by HPLC
revealed several obvious differences. The most apparent ones noted were
the absence of chlorophyll 12 and the associated decreased level of
chlorophyll A, a decreased level of neoxanthin and an increased
concentration of violaxanthin in extracts of K0 -9. The level of these two
xanthophylls appear to show an inverse relationship with neoxanthin
decreasing by approximately 70% and violaxanthin increasing by a
comparable amount. This noted relationship suggests that violaxanthin, a
presumed precursor to neoxanthin, undergoes only limited conversion in
IC0-9. Previous mutant studies on chlorophyll 12-deficient phenotypes of
algae and higher plants also noted decreased levels of neoxanthin and in
vitro reconstitution studies with the apoproteins of LHC-II showed that
rebinding of chlorophyll h required various xanthophylls, specifically
neoxanthin, also to be present.
The other light-harvesting complex mutants of Scenedesmus
examined in this study, including LF-1-LHC, LF-15 hf-17-LHC, CP-13-
LHC, PS-28-LHC and LF-23-LHC, exhibit similar general deficiencies of
chlorophyll h, lack the green bands of the LHCP-II but retain the
apoproteins of this complex and show the same relationship between
neoxanthin and violaxanthin as noted for K0 -9. Even though these strains
were derived from mutant phenotypes already suffering major deletions in
other parts of the photosynthetic apparatus, their inabilities to synthesize
or to accumulate chlorophyll J show the same common factor as noted for
K0 -9, i.e., the apparent accumulation of violaxanthin and decrease of
neoxanthin. This suggests that the mutual site of mutation in these
various phenotypes may reside at the point of conversion of violaxanthin to
neoxanthin. This interpretation supports the concept that the precursor(s) to chlorophyll b must be initially bound near to or at the site within the
chloroplast membrane where it will eventually function in the LHCP.
Examination by detergent polyacrylamide gel electrophoresis
(PAGE) of the light-harvesting complexes of isolated chloroplast fragments
of the several phenotypes employed in this investigation confirmed that the
inability to synthesize chlorophyll b was paralleled by the loss of the highly
pigmented LHCP-II. However, in no case was it observed that the
synthesis of the apoproteins of LHC-II, or their binding to the chloroplast
membrane, was affected by the mutation causing chlorophyll b deficiency.
Also no changes in electrophoretic mobility, i.e., apparent molecular
weight, of either the 26 or 28 kDa polypeptides were noted in electrophoretic
studies on control or heat denatured chloroplast fragments of the various
phenotypes. Similarly PAGE of chromatographically purified samples of
the LHC apoproteins of the WT and several mutants showed no alteration
of electrophoretic mobility. These observations suggest, but do not prove,
that the LHC-II apoproteins of the various phenotypes studied are not
modified by the mutation causing loss of chlorophyll h synthesis.
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