Biology Reference
In-Depth Information
Since in mixed PC/cholesterol membranes Ca
2
þ
ATPase activity exactly followed PC content
and was independent of cholesterol, they proposed that cholesterol was excluded from the
annulus (
Figure 10.18
). This very interesting concept was later shown via binding assays
to not be true. Later work from A.G. Lee and co-workers
[44,45,48,49]
confirmed the size
of the annular lipid ring, reporting a minimum number of annular phospholipid sites of
32 and 22 at 0
C and 37
C, respectively. Lee also extended the lipid/ Ca
2
þ
ATPase interaction
studies and found that anionic phospholipids PA, PS, and PI activate the protein while
binding at the general annular lipid sites and are involved in binding of Mg
2
þ
ATP. In addi-
tion, Ca
2
þ
ATPase activity is modified by bilayer acyl chain length, reaching a maximum at
C-18 and falling off at C-14 and C-22.
Protein Kinase C
Perhaps the most thoroughly studied membrane protein is protein kinase C (PKC), a key
enzyme in signal transduction
[50]
. The enzyme was only discovered by Yasutomi Nishizuka
in the late 1970s
[51]
. Phosphatidylserine (PS) is an essential cofactor that specifically binds to
and activates PKC. In fact all of the numerous isoforms of PKC are strictly dependent on PS
for activity. PKC specifically recognizes 1,2-sn-phosphatidyl-L-serine (the D isomer is ineffec-
tive)
[52]
. This and the fact that PS is not believed to be involved in cell signaling through the
formation of active metabolites, as is the case with PI, implies that specific, non-annular co-
factor binding sites for PS must exist on PKC. Also, PKC's specificity for PS requires diacyl-
glycerol. In the absence of diacylglycerol PKC binds anionic membranes with no discrimina-
tion between phospholipid headgroups beyond requiring a negative charge. PKC activity is
further modulated by bulk bilayer lipids related to curvature strain (discussed below). The
absolute specificity for PS is remarkable for known lipid
protein interactions.
e
Na
þ
/K
þ
ATPase
The plasma membrane-bound Na
þ
/K
þ
ATPase is responsible for maintaining essential
trans-membrane Na
þ
and K
þ
ion gradients. Its mechanism of action is discussed in Chapter
14. The Na
þ
/K
þ
ATPase was first described by Jens Skou (
Figure 10.20
) in 1957
[53]
, for
which he was awarded a 1997 Nobel Prize in Chemistry. The enzyme, isolated from Squalus
acanthus, was shown to bind 60 phospholipid molecules, comprising the annulus, with a pref-
erence for negatively charged lipids (K
3.8 for CL/PC, 1.7 for PS/PC, and 1.5 for PA/PC).
The effect of various lipids on stabilization of the Na
þ
,K
þ
-ATPase activity has been reported
by Haim et al.
[54]
. These authors reported that acidic phospholipids are required and PS is
somewhat better than PI. Also, optimal stabilization is achieved with heteroacid PSs having
a saturated fatty acid (18:0
¼
>
18:2) in the sn-
2 chain. In addition they also hypothesized that PS and cholesterol interact specifically with
each other near the
16:0) in the sn-1 and unsaturated fatty acid (18:1
b
1
subunit interface, thus stabilizing the protein. A general role for
anionic lipids at protein
a
1
/
protein interfaces is becoming more evident.
e
Potassium Channel KcsA
Recently Marius et al.
[55]
investigated the absolute requirement of anionic lipids for
potassium channel KcsA activity. The protein was isolated from Streptomyces lividans. High-
resolution X-ray crystallography showed an anionic lipid molecule bound to each of the
four protein
protein interfaces comprising the channel. In addition to the annular or
e
