Biology Reference
In-Depth Information
17.4.1 Fluorescence Properties of Chl and Freshly
Isolated Chloroplast at 77 K
Fluorescence spectroscopy at 77 K has been used extensively to probe the effects of
various metabolic tetrapyrroles on the state of organization of the chloroplast
membranes. It was deemed important therefore, to discuss the 77 K fluorescence
properties of Chl and freshly isolated chloroplasts before proceeding with a discus-
sion of experimental results.
17.4.1.1 Fluorescence Properties of Chlorophyll at 77 K
in Organic Solvents
Most of the light energy absorbed by Chls dissolved in organic solvents, is
dissipated as fluorescence. At the temperature of liquid N
2
(77 K) MV Chl
a
dissolved in diethyl ether coordinates to two solvent molecules (i.e. the central
Mg atom becomes hexacoordinated by axial coordination to two Lewis bases)
(Belanger and Rebeiz
1984
; Rebeiz and Belanger
1984
). It exhibits a major red
emission maximum at 674 nm [Qy (0
0
-0) transition], a minor maximum at 725 nm
[Qy (0
0
-1) transition], and Soret excitation maxima at 447 nm [By, Bx (0-0
0
)
transition] (Belanger and Rebeiz
1984
; Rebeiz and Belanger
1984
). It also exhibits,
a 422 nm (eta
η
2
transition) (Weiss
1975
,
1978
).
Under the same conditions, MV Chl
b
is also hexacoordinated and exhibits a major
red emission maximum at 659 nm [Qy (0
0
-0) transition], a minor maximum at
722 nm [Qy (0
0
-1) transition], and Soret excitation maxima at 475 nm [By (0-0
0
)
transition], 449 nm (
η
1
transition) and a 400 nm (eta
η
1
transition) and 427 nm (
η
2
transition) (Duggan and Rebeiz
1982
). The eta (
) transitions are forbidden in unsubstituted porphyrins (Weiss
1975
,
1978
), but become allowed in reduced porphyrins or when there is a conju-
gated carbonyl substituent as in the Chls (Weiss
1978
).
η
17.4.1.2 Fluorescence Properties of Chloroplasts at 77 K
In the chloroplast, MV Chl
a
and
b
are non-covalently associated with various
thylakoid polypeptides. This special pigment-protein environment changes drasti-
cally the population and energy levels of various electronic transitions and results in
different spectroscopic properties than in ether. As a consequence the spectroscopic
properties of a given Chl-polypeptide complex, depends on the specific Chl-protein
interactions within the complex. This picture is complicated further by the fact that
not all Chl-protein complexes are capable of fluorescence. Depending on the
structural proximity of various complexes, some Chl-polypeptides transfer their
excitation energy to other fluorescing complexes, instead of emitting their excitation
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