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<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>Dear Colleague, </span></font></p>
<p class=MsoNormal style='text-autospace:none'><font size=2 color=navy
face=Arial><span lang=EN-GB style='font-size:10.0pt;font-family:Arial;
color:navy'>I am sending a reference where I used sedimentation velocity in H2O
and D2O to evaluate partial specific volume and discuss the errors: </span></font></p>
<p class=MsoNormal style='text-autospace:none'><font size=2 color=navy
face=Arial><span lang=EN-GB style='font-size:10.0pt;font-family:Arial;
color:navy'>Gohon et al. </span></font><font size=2 face=Arial><span
lang=EN-GB style='font-size:10.0pt;font-family:Arial'>Analytical Biochemistry
334 (2004) 318–334</span></font></p>
<p class=MsoNormal style='text-autospace:none'><font size=2 face=Arial><span
lang=EN-GB style='font-size:10.0pt;font-family:Arial'>Please, if you want the
pdf file, send me an E mail. </span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>Using this method, the
precision is high when partial specific volume values are close to 1, (the
solvent density), i.e. for detergent, lipids...</span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>The precision is low of
for vbar values far from 1, i.e. for nucleic acids, charged polysaccharide...</span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>The method can be used to
heterogeneous solutions.</span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>The exchange of Hydrogen
to deuterium; and in the case of polyelectrolyte ion dissociation, have to be
considered in the calculation of partial specific volume.</span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>They affect in a small or
large extent the value of partial specific volume depending on the value of the
partial specific volume.</span></font></p>
<p class=MsoNormal><font size=2 color=navy face=Arial><span lang=EN-GB
style='font-size:10.0pt;font-family:Arial;color:navy'>All the best</span></font></p>
<p class=MsoNormal style='margin-bottom:12.0pt'><font size=2 color=navy
face=Arial><span lang=EN-GB style='font-size:10.0pt;font-family:Arial;
color:navy'>Christine</span></font></p>
<div>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span lang=EN-GB
style='font-size:10.0pt;font-family:"Courier New";color:navy'>Christine EBEL</span></font></p>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span
style='font-size:10.0pt;font-family:"Courier New";color:navy'>Institut de
Biologie Structurale CEA-CNRS-UJF</span></font></p>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span
style='font-size:10.0pt;font-family:"Courier New";color:navy'>41 rue Jules
Horowitz, F-38027 Grenoble France</span></font></p>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span
style='font-size:10.0pt;font-family:"Courier New";color:navy'>Tel (33) (0) 4 38
78 95 70; Fax (33) (0) 4 38 78 54 94 </span></font></p>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span
style='font-size:10.0pt;font-family:"Courier New";color:navy'>christine.ebel@ibs.fr</span></font></p>
<p class=MsoNormal><font size=2 color=navy face="Courier New"><span lang=EN-GB
style='font-size:10.0pt;font-family:"Courier New";color:navy'><a
href="http://www.ibs.fr/content/ibs_eng/presentation/lab/lbm/ebel.htm"><font
color=navy><span lang=FR style='color:navy'>http://www.ibs.fr/content/ibs_eng/presentation/lab/lbm/ebel.htm</span></font></a></span></font></p>
</div>
<p class=MsoNormal style='margin-left:35.4pt'><font size=2 face=Tahoma><span
style='font-size:10.0pt;font-family:Tahoma'>-----Message d'origine-----<br>
<b><span style='font-weight:bold'>De :</span></b> rasmb-bounces@rasmb.bbri.org
[mailto:rasmb-bounces@rasmb.bbri.org] <b><span style='font-weight:bold'>De la
part de</span></b> Ariel Lustig<br>
<b><span style='font-weight:bold'>Envoyé :</span></b> dimanche 8 juin 2008
14:03<br>
<b><span style='font-weight:bold'>À :</span></b> rasmb server1 forum<br>
<b><span style='font-weight:bold'>Objet :</span></b> [RASMB] Sv and Se for
V</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3
face="Times New Roman"><span style='font-size:12.0pt'> </span></font></p>
<div>
<p class=MsoNormal style='margin-left:35.4pt'><font size=2 face=Arial><span
lang=EN-GB style='font-size:10.0pt;font-family:Arial'>Dear Colleauges,</span></font></p>
</div>
<div>
<p class=MsoNormal style='margin-left:35.4pt'><font size=2 face=Arial><span
lang=EN-GB style='font-size:10.0pt;font-family:Arial'>I wanted to leave the
discussion (see Walter Staffords last e-mail 7 June 08) for the Newcastle
AUC Symposium in September 08, but it developed already now, so I
shall give here the abstract of the my </span></font></p>
</div>
<div>
<p class=MsoNormal style='margin-left:35.4pt'><font size=2 face=Arial><span
lang=EN-GB style='font-size:10.0pt;font-family:Arial'>contribution about
this theme and a new approach to solve it.:</span></font></p>
</div>
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<p class=MsoNormal style='margin-left:35.4pt'><font size=3
face="Times New Roman"><span lang=EN-GB style='font-size:12.0pt'> </span></font></p>
</div>
<div>
<p class=MsoNormal style='margin-left:35.4pt'><b><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial;font-weight:bold'>The use
of an elder mode to determine the density of vesicles by S</span></font></b><b><font
face="WP Greek Courier"><span lang=EN-GB style='font-family:"WP Greek Courier";
font-weight:bold'>0</span></font></b><b><font face=Arial><span lang=EN-GB
style='font-family:Arial;font-weight:bold'> versus ρ is a good approach that
can be used also to determine protein/RNA or protein/DNA complexes</span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'>. </span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>First of all we have to
bear in our mind that the molecular masses achieved from centrifugation, based
on the buoyant term (the partial specific volume,<b><span style='font-weight:
bold'> V </span></b>the particle), here in this discussed case <b><span
style='font-weight:bold'>V</span></b></span></font><b><font size=1 face=Arial><span
lang=EN-GB style='font-size:8.0pt;font-family:Arial;font-weight:bold'>pot.DNA</span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'> or <b><span
style='font-weight:bold'>V</span></b></span></font><b><font size=1 face=Arial><span
lang=EN-GB style='font-size:8.0pt;font-family:Arial;font-weight:bold'>prot.RNA</span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'>. Usually we invest a big
effort on sophisticated methods to calculate molecular weights <b><span
style='font-weight:bold'>Mw</span></b>, but the lack of precise <b><span
style='font-weight:bold'>V </span></b>values is like a weak link of a chain. In
such mentioned case we are able to calculate the <b><span
style='font-weight:bold'>V</span></b> from the protein amino acids and nucleic
acids but the exact ratio of both components is usually unknown. </span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>The classical density
determination by pycnometry even with digital precise instruments needs the
knowledge of the total particle concentration of the solution that can be only
estimated.</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>The use of S.J. Edelstein
and H.K.Schachman’s mode, (JBC 196, vol.247 , 306-317) to make
parallel runs in <b><span style='font-weight:bold'>H</span></b></span></font><b><font
size=1 face=Arial><span lang=EN-GB style='font-size:9.0pt;font-family:Arial;
font-weight:bold'>2</span></font></b><b><font face=Arial><span lang=EN-GB
style='font-family:Arial;font-weight:bold'>O</span></font></b><font face=Arial><span
lang=EN-GB style='font-family:Arial'> and <b><span style='font-weight:bold'>D</span></b></span></font><b><font
size=1 face=Arial><span lang=EN-GB style='font-size:9.0pt;font-family:Arial;
font-weight:bold'>2</span></font></b><b><font face=Arial><span lang=EN-GB
style='font-family:Arial;font-weight:bold'>O </span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'>buffer as mentioned also
in the RSC published book >> Analytical Ultracentrifugation,
Techniques and Methods<< page 197, edited by D. Scott et al,
can be employed only if the samples are pure, monodispersed and
reveal with sedimentation equilibrium SE straight slopes and single
Mw’s. The reality in case of biological samples is,
that the slopes tends to self-association , nonideality or
both effects and even show partly aggregation, so that the deviated
lines from them the V is calculated are not single and
straight.</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>Using the SEGAL computer
program when the slopes of 2 parts are calculated, it is <font
color=black><span style='color:black'>easy to realize this discrepancy.</span></font></span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>Using the <b><span
style='font-weight:bold'>S</span></b></span></font><b><font
face="WP Greek Courier"><span lang=EN-GB style='font-family:"WP Greek Courier";
font-weight:bold'>0</span></font></b><font face=Arial><span lang=EN-GB
style='font-family:Arial'> </span></font><b><font size=1 face=Arial><span
lang=EN-GB style='font-size:9.0pt;font-family:Arial;font-weight:bold'>relative</span></font></b><b><font
face=Arial><span lang=EN-GB style='font-family:Arial;font-weight:bold'> </span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'>versus<b><span
style='font-weight:bold'> ρ </span></b>mode<b><span style='font-weight:bold'> </span></b>developed
by C.H: Huang and J Charlton (JBC </span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>1971 Vol. 216) that was
used to study the density of vesicles, it is possible to apply it also for high
molecular complexes. The advantage in this case is, that even the sample
sediments with a boundary that contains partly a faster
or a slower component (as it happens often with biological samples) it
can be used to achieve <b><span style='font-weight:bold'>S°</span></b></span></font><b><font
face="WP Greek Courier"><span lang=EN-GB style='font-family:"WP Greek Courier";
font-weight:bold'>0</span></font></b><font face=Arial><span lang=EN-GB
style='font-family:Arial'> <b><span style='font-weight:bold'>(</span></b>at a D</span></font><font
size=1 face=Arial><span lang=EN-GB style='font-size:9.0pt;font-family:Arial'>2</span></font><font
face=Arial><span lang=EN-GB style='font-family:Arial'>O –H</span></font><font
size=1 face=Arial><span lang=EN-GB style='font-size:9.0pt;font-family:Arial'>2</span></font><font
face=Arial><span lang=EN-GB style='font-family:Arial'>O buffer density where S’
comes at rest) that represents the <b><span style='font-weight:
bold'>V</span></b> of the particle. Important is to use the whole series
of <b><span style='font-weight:bold'>S</span></b></span></font><b><font
face="WP Greek Courier"><span lang=EN-GB style='font-family:"WP Greek Courier";
font-weight:bold'>0</span></font></b><font face=Arial><span lang=EN-GB
style='font-family:Arial'> </span></font><b><font size=1 face=Arial><span
lang=EN-GB style='font-size:9.0pt;font-family:Arial;font-weight:bold'>relative </span></font></b><font
face=Arial><span lang=EN-GB style='font-family:Arial'>at the same particle
concentration to avoid S’ differences that stem from concentration dependence
of S’.</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>All our measurements were
done with absorption optics using the XLA at the same
wavelength <b><span style='font-weight:bold'>WL</span></b> since the used
samples do not show a distinct WL maximum at 260 nm or 280 nm.</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial'>...............yours......ariel</span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3
face="Times New Roman"><span style='font-size:12.0pt'> </span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><font size=3
face="Times New Roman"><span style='font-size:12.0pt'> </span></font></p>
<p class=MsoNormal style='margin-left:35.4pt'><b><font size=3 face=Arial><span
lang=EN-GB style='font-size:12.0pt;font-family:Arial;font-weight:bold'> </span></font></b></p>
<p class=MsoNormal align=center style='margin-left:35.4pt;text-align:center'><font
size=1 face=Arial><span lang=EN-GB style='font-size:9.0pt;font-family:Arial'> </span></font></p>
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