Re: Lys problem
ABRF (abrf@its.caltech.edu)
Tue, 21 Sep 1999 09:57:48 -0700
At 6:19 AM -0700 9/21/99, Pawel Mak wrote:
>Dear Colleagues,
>I have a question for protein sequencing experts: what may be cause of
>disappearing of lysine in sequenced samples? I use Applied Biosystems 491
>system and all is working perfectly: the instrument sequences samples very
>well, the level of lysine in standards is about 1.5 greater than level of
>leucine but mysteriously the potential lysine residue in sequenced proteins
>gives a blank run (in fact I can see lysine but it is very, very small peak
>and additionally before peC as well as after V appear small, unidentified
>peaks). Of course I know that the oxidants in A or in R4 can partially
>oxidise lysine but PARTIALLY... I also do not know if the very high level
>of DPU I observe on all chromatograms may be related to lysine problems ?
>Thank you very much for any suggestions !!!!
>Pawel Mak
>
>Dr. Pawel Mak
The conversion of DPTU to DPU is, of course, due to oxidation. PTH-Lysine
is very susceptible to oxidation as it contains two equivalents of
phenylthiocarbamyl. We don't have a 49X machine, but this problem
generally arises from voids in the system, particularly the entrance of the
delivery tube to the reaction cartridge. If this line is flush, then you
might begin to look for leaks around valve blocks or consider replacing old
teflon tubing. Best of luck.
Gary Hathaway
Gary M. Hathaway
Director, Protein Analytical Lab
Caltech, 139-74
400 S. Wilson Ave.
Pasadena, CA 91125
hathaway@cco.caltech.edu