PMID- 9814855
OWN - NLM
STAT- MEDLINE
DCOM- 19981123
LR  - 20190610
IS  - 0006-3002 (Print)
IS  - 0006-3002 (Linking)
VI  - 1374
IP  - 1-2
DP  - 1998 Sep 23
TI  - Agonist binding and affinity state transitions in reconstituted nicotinic 
      acetylcholine receptors revealed by single and sequential mixing stopped-flow 
      fluorescence spectroscopies.
PG  - 83-93
AB  - The affinity state of nicotinic acetylcholine receptors (nAcChoRs) reconstituted 
      into either dioleoylphosphatidylcholine (DOPC) or a mixture of 
      dioleoylphosphatidylcholine, dioleoylphosphatidic acid, and cholesterol 
      (DOPC/DOPA/cholesterol) has been determined using single and sequential mixing 
      stopped-flow fluorescence spectroscopies. These techniques have millisecond 
      temporal resolution, permitting low- and high-affinity conformational states of 
      the nAcChoR to be resolved following mixing with the fluorescent partial agonist 
      Dns-C6-Cho from their characteristic Dns-C6-Cho dissociation rates. Our studies 
      reveal that prior to agonist-induced affinity state conversion, nAcChoRs 
      reconstituted into either DOPC or DOPC/DOPA/cholesterol are predominantly in a 
      conformational state that has a low affinity for agonist. Prolonged exposure to 
      Dns-C6-Cho converts nearly all DOPC/DOPA/cholesterol-reconstituted nAcChoRs to 
      the high-affinity state. In contrast, Dns-C6-Cho converts only half of all 
      DOPC-reconstituted nAcChoRs to the high-affinity state. The other half persists 
      in a low-affinity state characterized by a Kd for Dns-C6-Cho of 0.61+/-0.07 
      microM. This Kd is similar to that previously reported for Dns-C6-Cho binding to 
      low-affinity, resting-state nAcChoRs in native membranes. However, affinity state 
      conversion of DOPC-reconstituted nAcChoRs may be facilitated by re-reconstituting 
      them into bilayers composed of DOPC/DOPA/cholesterol. These results indicate that 
      the lipid bilayer composition modulates nAcChoR agonist-induced affinity state 
      transitions.
FAU - Raines, D E
AU  - Raines DE
AD  - Department of Anesthesia and Critical Care, Massachusetts General Hospital, 
      Harvard Medical School, Boston 02114, USA. raines@etherdome.mgh.harvard.edu
FAU - Krishnan, N S
AU  - Krishnan NS
LA  - eng
GR  - GM53481/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - Netherlands
TA  - Biochim Biophys Acta
JT  - Biochimica et biophysica acta
JID - 0217513
RN  - 0 (Lipid Bilayers)
RN  - 0 (Nicotinic Agonists)
RN  - 0 (Phosphatidic Acids)
RN  - 0 (Phosphatidylcholines)
RN  - 0 (Receptors, Nicotinic)
RN  - 14268-17-8 (dioleoylphosphatidic acid)
RN  - 97C5T2UQ7J (Cholesterol)
RN  - EDS2L3ODLV (1,2-oleoylphosphatidylcholine)
SB  - IM
MH  - Animals
MH  - Binding Sites
MH  - Cholesterol/chemistry
MH  - Electric Organ/metabolism
MH  - In Vitro Techniques
MH  - Kinetics
MH  - Lipid Bilayers/chemistry
MH  - Nicotinic Agonists/*metabolism
MH  - Phosphatidic Acids/chemistry
MH  - Phosphatidylcholines/chemistry
MH  - Protein Conformation
MH  - Receptors, Nicotinic/*chemistry/*metabolism
MH  - Spectrometry, Fluorescence/*methods
MH  - Torpedo/metabolism
EDAT- 1998/11/14 00:00
MHDA- 1998/11/14 00:01
CRDT- 1998/11/14 00:00
PHST- 1998/11/14 00:00 [pubmed]
PHST- 1998/11/14 00:01 [medline]
PHST- 1998/11/14 00:00 [entrez]
AID - S0005-2736(98)00133-3 [pii]
AID - 10.1016/s0005-2736(98)00133-3 [doi]
PST - ppublish
SO  - Biochim Biophys Acta. 1998 Sep 23;1374(1-2):83-93. doi: 
      10.1016/s0005-2736(98)00133-3.