PMID- 10211825
OWN - NLM
STAT- MEDLINE
DCOM- 19990714
LR  - 20181113
IS  - 0961-8368 (Print)
IS  - 0961-8368 (Linking)
VI  - 8
IP  - 4
DP  - 1999 Apr
TI  - Helicity of alpha(404-451) and beta(394-445) tubulin C-terminal recombinant
      peptides.
PG  - 788-99
AB  - We have investigated the solution conformation of the functionally relevant
      C-terminal extremes of alpha- and beta-tubulin, employing the model recombinant
      peptides RL52alpha3 and RL33beta6, which correspond to the amino acid sequences
      404-451(end) and 394-445(end) of the main vertebrate isotypes of alpha- and
      beta-tubulin, respectively, and synthetic peptides with the
      alpha-tubulin(430-443) and beta-tubulin(412-431) internal sequences.
      Alpha(404-451) and beta(394-445) are monomeric in neutral aqueous solution (as
      indicated by sedimentation equilibrium), and have circular dichroism (CD) spectra
      characteristic of nearly disordered conformation, consistent with low scores in
      peptide helicity prediction. Limited proteolysis of beta(394-445) with
      subtilisin, instead of giving extensive degradation, resulted in main cleavages
      at positions Thr409-Glu410 and Tyr422-Gln423-Gln424, defining the proteolysis
      resistant segment 410-422, which corresponds to the central part of the predicted
      beta-tubulin C-terminal helix. Both recombinant peptides inhibited microtubule
      assembly, probably due to sequestration of the microtubule stabilizing associated
      proteins. Trifluoroethanol (TFE)-induced markedly helical CD spectra in
      alpha(404-451) and beta(394-445). A substantial part of the helicity of
      beta(394-445) was found to be in the CD spectrum of the shorter peptide
      beta(412-431) with TFE. Two-dimensional 1H-NMR parameters (nonsequential nuclear 
      Overhauser effects (NOE) and conformational C alphaH shifts) in 30% TFE permitted
      to conclude that about 25% of alpha(404-451) and 40% of beta(394-451) form
      well-defined helices encompassing residues 418-432 and 408-431, respectively,
      flanked by disordered N- and C-segments. The side chains of beta(394-451)
      residues Leu418, Val419, Ser420, Tyr422, Tyr425, and Gln426 are well defined in
      structure calculations from the NOE distance constraints. The apolar faces of the
      helix in both alpha and beta chains share a characteristic sequence of conserved 
      residues Ala,Met(+4),Leu(+7),Tyr(+11). The helical segment of alpha(404-451) is
      the same as that described in the electron crystallographic model structure of
      alphabeta-tubulin, while in beta(394-451) it extends for nine residues more,
      supporting the possibility of a functional coil --> helix transition at the
      C-terminus of beta-tubulin. These peptides may be employed to construct model
      complexes with microtubule associated protein binding sites.
FAU - Jimenez, M A
AU  - Jimenez MA
AD  - Instituto de Estructura de la Materia, CSIC, Madrid, Spain.
FAU - Evangelio, J A
AU  - Evangelio JA
FAU - Aranda, C
AU  - Aranda C
FAU - Lopez-Brauet, A
AU  - Lopez-Brauet A
FAU - Andreu, D
AU  - Andreu D
FAU - Rico, M
AU  - Rico M
FAU - Lagos, R
AU  - Lagos R
FAU - Andreu, J M
AU  - Andreu JM
FAU - Monasterio, O
AU  - Monasterio O
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Protein Sci
JT  - Protein science : a publication of the Protein Society
JID - 9211750
RN  - 0 (Recombinant Proteins)
RN  - 0 (Tubulin)
RN  - 75-89-8 (Trifluoroethanol)
RN  - J41CSQ7QDS (Zinc)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Chickens
MH  - Chromatography, High Pressure Liquid
MH  - Circular Dichroism
MH  - Computer Simulation
MH  - Humans
MH  - Magnetic Resonance Spectroscopy
MH  - Models, Molecular
MH  - Molecular Sequence Data
MH  - Plasmids
MH  - *Protein Structure, Secondary
MH  - Recombinant Proteins/chemistry
MH  - Time Factors
MH  - Trifluoroethanol/chemistry
MH  - Tubulin/*chemistry
MH  - Ultracentrifugation
MH  - Zinc/chemistry
PMC - PMC2144297
EDAT- 1999/04/22 00:00
MHDA- 1999/04/22 00:01
CRDT- 1999/04/22 00:00
PHST- 1999/04/22 00:00 [pubmed]
PHST- 1999/04/22 00:01 [medline]
PHST- 1999/04/22 00:00 [entrez]
AID - 10.1110/ps.8.4.788 [doi]
PST - ppublish
SO  - Protein Sci. 1999 Apr;8(4):788-99. doi: 10.1110/ps.8.4.788.