If you have a very pure protein sample, you'll want to use an ESI-ion trap for analyzing proteins of that size. It should be possible to get an exact mass (i.e. within a single Da). It's possible, but very rare, to get exact masses of proteins up to 100 kDa using ESI-ion trap instruments.
If your sample is not highly purified, you'll need to use some type of TOF instrument. MALDI-TOF should work. With a TOF instrument, you shouldn't expect to be able to distinguish between point mutants of a protein, but you'll be able to get information about larger changes. For instance, you'll be able to determine if you completely cleaved a his-tag off a construct.
Our lab is spoiled because we have access to the HHMI Mass Spec facility, but I'd imagine that there are many facilities that can get accurate masses on 40 kDa proteins.
-Joel
=================================================
Joel M. Guenther
PhD Candidate, Department of Chemistry
Kuriyan Laboratory
http://jkweb.berkeley.edu/
University of California, Berkeley
176 Stanley Hall, QB3
Berkeley, CA 94720-3220
tel: (510) 643 0166
fax: (510) 643 2352
=================================================
On 9/5/07, Jacob Keller <[log in to unmask]
> wrote:I second Dr. Loll's question, and would like to be CC'd in whatever MS tips, including
service-providers, are sent. I have been having a bit of a debacle with a certain MS service provider.
Jacob Keller
==============Original message text===============
On Wed, 05 Sep 2007 11:41:52 am CDT Patrick Loll wrote:
I wonder if anyone would care to share experiences/ideas/biases that
relate to the use of mass spectrometry to verify the identity of
protein constructs used for crystallization. Our experience with
different MS facilities has been checquered.
Specifically:
What's the current thinking on the best approach to get masses for
intact proteins of moderate size (say, 40 kD)? ESI-TOF?
What kind of resolution should one hope to obtain in such cases
(10E-04?)
Any suggestions as to good facilities offering fee for service MS
characterization are welcome (but should be shared off line, I think;
continental US only).
Thanks,
Pat
------------------------------------------------------------------------
---------------
Patrick J. Loll, Ph. D. (215) 762-7706
Associate Professor FAX: (215) 762-4452
Department of Biochemistry & Molecular Biology
Director, Biochemistry Graduate Program
Drexel University College of Medicine
Room 10-102 New College Building
245 N. 15th St., Mailstop 497
Philadelphia, PA 19102-1192 USA
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===========End of original message text===========
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Jacob Keller
Northwestern University
6541 N. Francisco #3
Chicago IL 60645
(847)491-2438
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