TY - JOUR
T1 - Frequency-domain measurements of the rotational dynamics of the tyrosine groups of calmodulin
AU - Gryczynski, I.
AU - Lakowicz, J. R.
AU - Steiner, R. F.
N1 - Funding Information:
conformationalc hange,w hich reducest he separation of the tyrosines and increasest he efficiency of transfer.T he tertiarys tructureso f the fragmentsa nd hence the microenvironmentos f Tyr-99 and Tyr-138a ppeart o be similart o that of nativec almodulin[ 21]. The anisotropyd ecay parametersg iven in tables 3 and 4 werec omputedu singt he fixed values of r,, obtained by direct measuremenitn 70% glycerol at - 60°C. Since the energyt ransferi s expected to persist at low temperaturest hese parametersp robably reflect dominatelyt he motional propertieso f the residuesa, nd not the com-ponentd ue to energyt ransferW. e believet he time scale of the transfer may be too rapid to be recoveredf rom the present data, and that the processs ervesp rimarily to reduce the apparent value of r,. It is well known that rapid components in an anisotropy decay, which are faster than the time resolution of the measurements, result in decreaseda pparentv aluesf or r, [22,23]. This suggestion is supported by an alternative analysiso f the anisotropyd ata,i n which Q was a
Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.
PY - 1988/5
Y1 - 1988/5
N2 - We used frequency-domain fluorometry to determine the intensity and anisotropy decay kinetics of tyrosine residues in calmodulin and its fragments. Excitation was provided by a continuous ultraviolet laser source, a frequency-doubled rhodamine 6G ring dye laser, whose output was externally modulated to 200 MHz. Both the intensity and anisotropy decays were found to be multiexponential and dependent upon temperature and solution conditions. By examination of calmodulin fragments we determined that energy transfer between the two tyrosine residues reduces the steady-state anisotropy values by about 20%. Additionally, the frequency-domain anisotropy decays indicate local torsional motions of the tyrosine residues, as well as significant individual motions of the two domains of calmodulin.
AB - We used frequency-domain fluorometry to determine the intensity and anisotropy decay kinetics of tyrosine residues in calmodulin and its fragments. Excitation was provided by a continuous ultraviolet laser source, a frequency-doubled rhodamine 6G ring dye laser, whose output was externally modulated to 200 MHz. Both the intensity and anisotropy decays were found to be multiexponential and dependent upon temperature and solution conditions. By examination of calmodulin fragments we determined that energy transfer between the two tyrosine residues reduces the steady-state anisotropy values by about 20%. Additionally, the frequency-domain anisotropy decays indicate local torsional motions of the tyrosine residues, as well as significant individual motions of the two domains of calmodulin.
KW - Calmodulin
KW - Fluorescence anisotropy decay
KW - Fluorescence intensity decay
KW - Frequency-domain fluorometry
KW - Tyrosine
UR - http://www.scopus.com/inward/record.url?scp=0024009994&partnerID=8YFLogxK
U2 - 10.1016/0301-4622(88)85003-8
DO - 10.1016/0301-4622(88)85003-8
M3 - Article
C2 - 3416036
AN - SCOPUS:0024009994
SN - 0301-4622
VL - 30
SP - 49
EP - 59
JO - Biophysical Chemistry
JF - Biophysical Chemistry
IS - 1
ER -