PMID- 10354417
OWN - NLM
STAT- MEDLINE
DCOM- 19990712
LR  - 20231213
IS  - 0006-3495 (Print)
IS  - 1542-0086 (Electronic)
IS  - 0006-3495 (Linking)
VI  - 76
IP  - 6
DP  - 1999 Jun
TI  - The role of a conserved proline residue in mediating conformational changes 
      associated with voltage gating of Cx32 gap junctions.
PG  - 2887-98
AB  - We have explored the role of a proline residue located at position 87 in the 
      second transmembrane segment (TM2) of gap junctions in the mechanism of 
      voltage-dependent gating of connexin32 (Cx32). Substitution of this proline 
      (denoted Cx32P87) with residues G, A, or V affects channel function in a 
      progressive manner consistent with the expectation that a proline kink (PK) motif 
      exists in the second transmembrane segment (TM2) of this connexin. Mutations of 
      the preceding threonine residue T86 to S, A, C, V, N, or L shift the 
      conductance-voltage relation of wild-type Cx32, such that the mutated channels 
      close at smaller transjunctional voltages. The observed shift in voltage 
      dependence is consistent with a reduction in the open probability of the mutant 
      hemichannels at a transjunctional voltage (Vj) of 0 mV. In both cases in which 
      kinetics were examined, the time constants for reaching steady state were faster 
      for T86N and T86A than for wild type at comparable voltages, suggesting that the 
      T86 mutations cause the energetic destabilization of the open state relative to 
      the other states of the channel protein. The structural underpinnings of the 
      observed effects were explored with Monte Carlo simulations. The conformational 
      space of TM2 helices was found to differ for the T86A, V, N, and L mutants, which 
      produce a less bent helix ( approximately 20 degrees bend angle) compared to the 
      wild type, which has a approximately 37 degrees bend angle. The greater bend 
      angle of the wild-type helix reflects the propensity of the T86 residue to 
      hydrogen bond with the backbone carbonyl of amino acid residue I82. The relative 
      differences in propensity for hydrogen bonding of the mutants relative to the 
      wild-type threonine residue in the constructs we studied (T86A, V, N, L, S, and 
      C) correlate with the shift in the conductance-voltage relation observed for T86 
      mutations. The data are consistent with a structural model in which the open 
      conformation of the Cx32 channel corresponds to a more bent TM2 helix, and the 
      closed conformation corresponds to a less bent helix. We propose that the 
      modulation of the hydrogen-bonding potential of the T86 residue alters the bend 
      angle of the PK motif and mediates conformational changes between open and closed 
      channel states.
FAU - Ri, Y
AU  - Ri Y
AD  - Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York 
      10461, USA.
FAU - Ballesteros, J A
AU  - Ballesteros JA
FAU - Abrams, C K
AU  - Abrams CK
FAU - Oh, S
AU  - Oh S
FAU - Verselis, V K
AU  - Verselis VK
FAU - Weinstein, H
AU  - Weinstein H
FAU - Bargiello, T A
AU  - Bargiello TA
LA  - eng
GR  - R01 GM046889/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PL  - United States
TA  - Biophys J
JT  - Biophysical journal
JID - 0370626
RN  - 0 (Connexins)
RN  - 9DLQ4CIU6V (Proline)
SB  - IM
MH  - Animals
MH  - Biophysical Phenomena
MH  - Biophysics
MH  - Computer Simulation
MH  - Connexins/*chemistry/genetics/*metabolism
MH  - Electrophysiology
MH  - Female
MH  - Gap Junctions/*metabolism
MH  - Hydrogen Bonding
MH  - In Vitro Techniques
MH  - Membrane Potentials
MH  - Models, Molecular
MH  - Monte Carlo Method
MH  - Mutagenesis, Site-Directed
MH  - Oocytes/metabolism
MH  - Proline/chemistry
MH  - Protein Conformation
MH  - Protein Structure, Secondary
MH  - Xenopus
MH  - Gap Junction beta-1 Protein
PMC - PMC1300261
EDAT- 1999/06/04 00:00
MHDA- 1999/06/04 00:01
PMCR- 2000/06/01
CRDT- 1999/06/04 00:00
PHST- 1999/06/04 00:00 [pubmed]
PHST- 1999/06/04 00:01 [medline]
PHST- 1999/06/04 00:00 [entrez]
PHST- 2000/06/01 00:00 [pmc-release]
AID - S0006-3495(99)77444-8 [pii]
AID - 10.1016/S0006-3495(99)77444-8 [doi]
PST - ppublish
SO  - Biophys J. 1999 Jun;76(6):2887-98. doi: 10.1016/S0006-3495(99)77444-8.