<html dir="ltr">
<head>
<meta http-equiv="Content-Type" content="text/html; charset=iso-8859-1">
<style type="text/css" id="owaParaStyle"></style>
</head>
<body fpstyle="1" ocsi="0">
<div style="direction: ltr;font-family: Tahoma;color: #000000;font-size: 10pt;">
<div>Hi all,</div>
<div><br>
</div>
We are trying a new (?) method of determining dissociation constants for protein/DNA complexes using velocity sedimentation and I am looking for feedback regarding the method.
<div><br>
</div>
<div>Our method of determining Kds uses fluorescently-labeled oligos in complex with unlabeled protein. By scanning at 488 nm, we can determine the size distribution of both unbound DNA and the protein/DNA complex. We then calculate the area under each of
the (baseline resolved) distribution curves, normalize these values to 100% total, and use the ratios (along with our known initial concentrations) to calculate the molar concentrations of bound/unbound protein/DNA. From these data, we are able to get Kds
that correspond well to Kds calculated from other experiments. </div>
<div><br>
<div>We are looking to publish these results, and I was wondering if anyone knew of Kds calculated in this manner before (either in published literature or otherwise)? Any suggestions/comments welcome, thanks!</div>
<div><br>
</div>
<div>Best,</div>
<div>Sarah<br>
<div><br>
<div style="font-family:Tahoma; font-size:13px">
<div style="font-family:Tahoma; font-size:13px">___________________________
<div>Sarah Kessans</div>
<div>Post-doctoral Fellow</div>
<div>Biomolecular Interaction Centre</div>
<div>School of Biological Sciences</div>
<div>University of Canterbury</div>
</div>
</div>
</div>
</div>
</div>
</div>
</body>
</html>
<HTML><P><PRE>
This email may be confidential and subject to legal privilege, it may
not reflect the views of the University of Canterbury, and it is not
guaranteed to be virus free. If you are not an intended recipient,
please notify the sender immediately and erase all copies of the message
and any attachments.
Please refer to http://www.canterbury.ac.nz/emaildisclaimer for more
information.
</PRE></P></HTML>