PMID- 8662678
OWN - NLM
STAT- MEDLINE
DCOM- 19960815
LR  - 20210210
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 271
IP  - 22
DP  - 1996 May 31
TI  - Modification of the pH profile and tetrabenazine sensitivity of rat VMAT1 by 
      replacement of aspartate 404 with glutamate.
PG  - 13048-54
AB  - Vesicular monoamine transporters (VMAT) catalyze transport of serotonin, 
      dopamine, epinephrine, and norepinephrine into subcellular storage organelles in 
      a variety of cells. Accumulation of the neurotransmitter depends on the proton 
      electrochemical gradient (Delta micro H+) across the organelle membrane and 
      involves VMAT-mediated exchange of two lumenal protons with one cytoplasmic 
      amine. Mutagenic analysis of the role of two conserved Asp residues located in 
      transmembrane segments X and XI of rat VMAT type I reveals an important role of 
      these two residues in catalysis. Replacement of Asp 431 with either Glu or Ser 
      inhibits VMAT-mediated [3H]serotonin transport. The mutated proteins are 
      unimpaired in ligand recognition as measured with the high affinity ligand 
      [3H]reserpine or coupling to the proton electrochemical gradient as judged by its 
      ability to accelerate [3H]reserpine binding. Therefore, the Asp residue is needed 
      as such in this position and even a conservative replacement with Glu generates a 
      protein that can catalyze only partial reactions but cannot complete the 
      transport cycle. Replacement of Asp 404 with either Ser or Cys inhibits all 
      VMAT-mediated reactions measured. However, replacement with Glu generated a 
      protein that catalyzed [3H]serotonin transport with modified properties. Whereas 
      the mutated protein binds [3H]reserpine to normal levels and the pH optimum of 
      this reaction is only slightly affected, the optimum pH for transport activity 
      shifted to the acid side and became very sharp; in addition the sensitivity to 
      the inhibitor tetrabenazine increased significantly in this mutated protein. The 
      results point to the need of a carboxyl moiety in position 404. A slight change 
      in its relative location or in the environment around it has a significant effect 
      on the pK of group(s) involved in steps after ligand recognition and coupling to 
      the first H+.
FAU - Steiner-Mordoch, S
AU  - Steiner-Mordoch S
AD  - Alexander Silberman Institute of Life Sciences, Hebrew University, Jerusalem, 
      91904 Israel.
FAU - Shirvan, A
AU  - Shirvan A
FAU - Schuldiner, S
AU  - Schuldiner S
LA  - eng
GR  - NS16708/NS/NINDS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (Membrane Glycoproteins)
RN  - 0 (Membrane Transport Proteins)
RN  - 0 (Neuropeptides)
RN  - 0 (Slc18a1 protein, rat)
RN  - 0 (Vesicular Biogenic Amine Transport Proteins)
RN  - 0 (Vesicular Monoamine Transport Proteins)
RN  - 10028-17-8 (Tritium)
RN  - 30KYC7MIAI (Aspartic Acid)
RN  - 333DO1RDJY (Serotonin)
RN  - 3KX376GY7L (Glutamic Acid)
RN  - Z9O08YRN8O (Tetrabenazine)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Aspartic Acid/*genetics
MH  - Base Sequence
MH  - Biological Transport
MH  - Catalysis
MH  - Cell Line
MH  - Glutamic Acid/*genetics
MH  - Hydrogen-Ion Concentration
MH  - Membrane Glycoproteins/*drug effects/genetics/metabolism
MH  - *Membrane Transport Proteins
MH  - Molecular Sequence Data
MH  - Mutagenesis, Site-Directed
MH  - *Neuropeptides
MH  - Rats
MH  - Serotonin/metabolism
MH  - Tetrabenazine/*pharmacology
MH  - Tritium
MH  - Vesicular Biogenic Amine Transport Proteins
MH  - Vesicular Monoamine Transport Proteins
EDAT- 1996/05/31 00:00
MHDA- 1996/05/31 00:01
CRDT- 1996/05/31 00:00
PHST- 1996/05/31 00:00 [pubmed]
PHST- 1996/05/31 00:01 [medline]
PHST- 1996/05/31 00:00 [entrez]
AID - S0021-9258(18)41470-6 [pii]
AID - 10.1074/jbc.271.22.13048 [doi]
PST - ppublish
SO  - J Biol Chem. 1996 May 31;271(22):13048-54. doi: 10.1074/jbc.271.22.13048.