Raines D E, Miller K W
Department of Anesthesia, Massachusetts General Hospital, Boston 02114.
Biophys J. 1993 Mar;64(3):632-41. doi: 10.1016/S0006-3495(93)81422-X.
The effect of salt and pH titration on the selectivity of spin-labeled analogues of phosphatidic acid, phosphatidylserine, phosphatidylcholine, and stearic acid for the nicotinic acetylcholine receptor (nAcChoR) reconstituted into dioleoylphosphatidylcholine was examined at 0 degrees C using electron spin resonance spectroscopy. The order of selectivity at pH 7.4 and 0 mM NaCl was phosphatidylserine > stearic acid > phosphatidic acid > phosphatidylcholine. The addition up to 2 M NaCl or titration of pH from 5.0 to > 9.0 did not alter the selectivity of the phospholipids for the nAcChoR. For stearic acid, conversely, titration of pH from 5.0 to 9.0 at 0 mM NaCl and titration of NaCl from 0 to 2 M at pH 9.0 both increased selectivity for the nAcChoR. It is concluded that electrostatic interactions do not account for the selectivity of the negatively charged phospholipids, phosphatidylserine, and phosphatidic acid for the nAcChoR. This is consistent with the known orientation of the transmembrane sequences M1 and M4, which predicts a balance in the number of negative and positive charges in the lipid-protein interface and suggests that the two positive charges on each M3 helix are not exposed to the lipid-protein interface.
在0℃下，使用电子自旋共振光谱法研究了盐和pH滴定对磷脂酸、磷脂酰丝氨酸、磷脂酰胆碱和硬脂酸的自旋标记类似物对重组入二油酰磷脂酰胆碱中的烟碱型乙酰胆碱受体（nAcChoR）的选择性的影响。在pH 7.4和0 mM NaCl条件下，选择性顺序为磷脂酰丝氨酸＞硬脂酸＞磷脂酸＞磷脂酰胆碱。添加高达2 M NaCl或将pH从5.0滴定至＞9.0不会改变磷脂对nAcChoR的选择性。相反，对于硬脂酸，在0 mM NaCl条件下将pH从5.0滴定至9.0以及在pH 9.0条件下将NaCl从0滴定至2 M均增加了对nAcChoR的选择性。得出的结论是，静电相互作用不能解释带负电荷的磷脂、磷脂酰丝氨酸和磷脂酸对nAcChoR的选择性。这与跨膜序列M1和M4的已知取向一致，该取向预测脂质-蛋白质界面中负电荷和正电荷数量的平衡，并表明每个M3螺旋上的两个正电荷不暴露于脂质-蛋白质界面。
