[ccp4bb] Thanks for the many responses

[Nicholas Keep]

Thanks for the many responses.  I had been setting the parameters right. The problem was I was not telling xds to go 
back as far as INIT in the reprocessing.  JOB ALL was the answer

Best wishes
Nick
--

Prof Nicholas H. Keep
Executive Dean of School of Science
Professor of Biomolecular Science
Crystallography, Institute for Structural and Molecular Biology,
Department of Biological Sciences
Birkbeck,  University of London,
Malet Street,
Bloomsbury
LONDON
WC1E 7HX

email n.k...@mail.cryst.bbk.ac.uk
Telephone 020-7631-6852  (Room G57 Office)
  020-7631-6800  (Department Office)
Fax   020-7631-6803
If you want to access me in person you have to come to the crystallography 
entrance
and ring me or the department office from the internal phone by the door

[ccp4bb] Thanks for the many responses concerning protein concentration

[Mark Hilge]

Dear colleagues,

Thanks a lot for the many constructive suggestions! Attchached is  
another response I got directly.


Best regards,

Mark

__

Hello Mark,

for low absorbing proteins, the Ehresmann method is a good choice. This
empiric formula converts absorption into protein concentration:

(A228.5-A234.5)/3.14 = [protein] in mg/mL

The method was developed to determine whole protein concentration in  
plant
extracts that also contain nucleic acid. These absorb the same at the  
two

wavelengths, so the difference is proportional to the protein
concentration provided there are no other absorbing agents (such as
thiols, old MOPS buffer etc.). 3.14 is an empiric conversion factor
unrelated to pi.

Since at these wavelengths the amide bonds absorb, one can get away with
very little protein. I have tried this in a nanodrop, but not
exhaustively. If the monochromator can not step with 0.5nm, four
wavelengths may be used as approximation.

Ref.:

Ehresmann B, Imbault P, Weil JH.
Spectrophotometric determination of protein concentration in cell  
extracts

containing tRNA's and rRNA's.
Anal Biochem. 1973 Aug;54(2):454-63.

HTH, regards,

Markus

__

Mark Hilge
Protein Biophysics
NCMLS 274
3rd floor M850.03.035
Geert Grooteplein 28
6525 GA Nijmegen
The Netherlands

http://www.mark-hilge.com

Phone: 0031 24 36 10 525



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