A comparative Raman spectroscopic study of cholinesterases.
We report Raman spectra of various cholinesterases: lytic tetrameric forms (G4) obtained by tryptic digestion of asymmetric acetylcholinesterase (AChE) from Torpedo californica and Electrophorus electricus, a PI-PLC-treated dimeric form (G2) of AChE from T marmorata, and the soluble tetrameric form (G4) of butyrylcholinesterase (BuChE) from human plasma. The contribution of different types of secondary structure was estimated by analyzing the amide I band, using the method of Williams. The spectra of cholinesterases in 10 mM Tris-HCl (pH 7.0) indicate the presence of both alpha-helices (about 50%) and beta-sheets (about 25%), together with 15% turns and 10% undefined structures. In 20 mM phosphate buffer (pH 7.0), the spectra indicated a smaller contribution of alpha-helical structure (about 35%) and an increased beta-sheet content (from 25 to 35%). This shows that the ionic milieu profoundly affects either the conformation of the protein (AChE activity is known to be sensitive to ionic strength), or the evaluation of secondary structure, or both. In addition, we analyzed vibrations corresponding to the side chains of aromatic and aliphatic amino acids. In particular, the analyses of the tyrosine doublet (830-850 cm-1) and of the tryptophan vibration at 880 cm-1 indicated that these residues are predominantly 'exposed' on the surface of the molecules.
Aslanian
D
D
Laboratoire de physique des solides, Associé au CNRS, Université P et M Curie, Paris, France.
Grof
P
P
Bon
S
S
Masson
P
P
Négrerie
M
M
Chatel
J M
JM
Balkanski
M
M
Taylor
P
P
Massoulié
J
J
D000110:Acetylcholinesterase / Q000737:chemistry* / Q000302:isolation & purification; D000818:Animals; D002091:Butyrylcholinesterase / Q000737:chemistry* / Q000302:isolation & purification; D004593:Electrophorus; D006801:Humans; D010649:Phenylalanine / Q000737:chemistry; D011487:Protein Conformation; D013059:Spectrum Analysis, Raman; D014101:Torpedo; D014364:Tryptophan / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry
D014443:Tyrosine; D010649:Phenylalanine; D014364:Tryptophan; D000110:Acetylcholinesterase; D002091:Butyrylcholinesterase