TY - JOUR
T1 - Association of tetramers of human butyrylcholinesterase is mediated by conserved aromatic residues of the carboxy terminus
AU - Altamirano, Cibby Varkey
AU - Lockridge, Oksana
N1 - Funding Information:
We thank Dr Cheng Kao (University of Indiana, Bloomington, IN) for two-hybrid system plasmids and yeast strain, and Dr Eric Krejci (Laboratoire de Neurobiologie Cellulaire et Moléculaire, Paris, France) for rat collagen tail (Q subunit) cDNA. This work was supported by US Medical Research and Material Command DAMD17-94-J-4005, DAMD17-97-1-7349, and AASERT award DAAG55-97-1-0244.
PY - 1999/5/14
Y1 - 1999/5/14
N2 - Human butyrylcholinesterase (BChE) is composed predominantly of tetramers. Our laboratory has shown that up to 40 carboxy terminal residues of each subunit contribute to the stabilization of tetramers (R.M. Blong, E. Bedows, O. Lockridge, The tetramerization domain of butyrylcholinesterase is at the carboxy-terminus, Biochem. J. 327 (1997) 747-757). To better define the residues which participate in tetramer stabilization, the in vivo interaction of the BChE C-terminus 46 residue peptide was quantitated for wild type and mutant BChE using the yeast two-hybrid system. The wild type C-terminal peptides interacted with one another in this system. The K-variant (A539T) and C571A peptides showed interaction similar to that of the wild type. However, only 11.7% of the interaction seen with the wild type peptide was observed with the mutant in which seven conserved aromatic residues (Trp 543, Phe 547, Trp 550, Tyr 553, Trp 557, Phe 561, and Tyr 564) had been altered to alanines (aromatics off mutant). When these seven mutations were incorporated into the complete BChE molecule and expressed in 293T cells, only monomers and dimers were observed. The addition of poly-l-proline to the medium of 293T cells expressing wild type BChE resulted in the increase of the tetrameric form, similar to that observed by Bon et al. (S. Bon, F. Coussen, J. Massoulie, Quaternary associations of acetylcholinesterase II. The polyproline attachment domain of the collagen tail, J. Biol. Chem. 272 (1997) 3016-3021) for acetylcholinesterase expressed in COS cells. However, no increase in tetramers was observed with poly-l-proline addition to the medium of 293T cells expressing the aromatics off BChE mutant. These observations suggest that the stabilization of BChE tetramers is mediated through the interaction of the seven conserved aromatic residues, Trp 543, Phe 547, Trp 550, Tyr 553, Trp 557, Phe 561, and Tyr 564, and that the poly-l-proline induced increase in tetrameric BChE is mediated through these seven aromatic residues. Copyright (C) 1999 Elsevier Science Ireland Ltd.
AB - Human butyrylcholinesterase (BChE) is composed predominantly of tetramers. Our laboratory has shown that up to 40 carboxy terminal residues of each subunit contribute to the stabilization of tetramers (R.M. Blong, E. Bedows, O. Lockridge, The tetramerization domain of butyrylcholinesterase is at the carboxy-terminus, Biochem. J. 327 (1997) 747-757). To better define the residues which participate in tetramer stabilization, the in vivo interaction of the BChE C-terminus 46 residue peptide was quantitated for wild type and mutant BChE using the yeast two-hybrid system. The wild type C-terminal peptides interacted with one another in this system. The K-variant (A539T) and C571A peptides showed interaction similar to that of the wild type. However, only 11.7% of the interaction seen with the wild type peptide was observed with the mutant in which seven conserved aromatic residues (Trp 543, Phe 547, Trp 550, Tyr 553, Trp 557, Phe 561, and Tyr 564) had been altered to alanines (aromatics off mutant). When these seven mutations were incorporated into the complete BChE molecule and expressed in 293T cells, only monomers and dimers were observed. The addition of poly-l-proline to the medium of 293T cells expressing wild type BChE resulted in the increase of the tetrameric form, similar to that observed by Bon et al. (S. Bon, F. Coussen, J. Massoulie, Quaternary associations of acetylcholinesterase II. The polyproline attachment domain of the collagen tail, J. Biol. Chem. 272 (1997) 3016-3021) for acetylcholinesterase expressed in COS cells. However, no increase in tetramers was observed with poly-l-proline addition to the medium of 293T cells expressing the aromatics off BChE mutant. These observations suggest that the stabilization of BChE tetramers is mediated through the interaction of the seven conserved aromatic residues, Trp 543, Phe 547, Trp 550, Tyr 553, Trp 557, Phe 561, and Tyr 564, and that the poly-l-proline induced increase in tetrameric BChE is mediated through these seven aromatic residues. Copyright (C) 1999 Elsevier Science Ireland Ltd.
KW - Butyrylcholinesterase
KW - K-variant
KW - Poly-L-proline
KW - Proline-rich attachment domain
KW - Tetramerization domain
KW - Yeast two-hybrid system
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U2 - 10.1016/S0009-2797(99)00013-7
DO - 10.1016/S0009-2797(99)00013-7
M3 - Article
C2 - 10421438
AN - SCOPUS:18744437671
VL - 119-120
SP - 53
EP - 60
JO - Chemico-Biological Interactions
JF - Chemico-Biological Interactions
SN - 0009-2797
ER -