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Re: kinases Protein kinase band shifts



>To those of you out there that have used Western blotting to look at the
>regulation of protein kinases, I would greatly appreciate your help. We
>have been developing a technique in my laboratory to monitor the regulation
>of over 50 different protein kinases at once on a single SDS-PAGE gel. The
>method exploits the bandshifts that often occurs when protein kinases and
>their substrates undergo phosphorylation. So far, in diverse model systems,
>we have detected band shifts in the following protein-serine kinases:
>cAMP-dependent protein kinase, cGMP-dependent protein kinase,
>PKBalpha(Akt1), PKBbeta(Akt2), PKCbeta, PKCzeta, PKCepsilon, Erk1, Erk2,
>Rsk1, Rsk2, p70 S6 kinase, p46 JNK(SAPK), p54 JNK(SAPK), Pak1, Mek4, Mek6
>and casein kinase 2. We have also seen very small bandshifts in p38alpha
>(Hog) MAP kinase. We have failed to see band-shifts in Mek1 and MAPKAPK2
>under conditions where these kinases should have been phosphorylated and
>activated.
>
>I would be very grateful to learn from any of you whether you are aware of
>any other protein-serine kinases or protein-tyrosine kinases that exhibit
>bandshifting on SDS-PAGE gels, the model system where the bandshift was
>seen and what stimulus was used. For those that send me an e-mail reply
>with such information, I will send back a list of the findings that members
>of this discussion group have provided. If enough people participate and
>the list is large, then I will transmit it to everyone (anonymity would be
>maintained).
>
>I believe that such band shift assays for kinase changes are an excellent
>way to probe whether kinases are regulated in different model systems.
>Kinases that display such changes can be further examined with
>phospho-specific antibodies or in phosphotransferase activity assays. If
>all goes well, then I may be able to provide such analysis as a service
>those in the signal transduction community.
>
>Thank you from Steven Pelech.
>
>Steven Pelech, Ph.D.
>Professor
>Department of Medicine
>Rm S125, 2nd Floor, Koerner Pavilion
>2211 Wesbrook Mall
>University of British Columbia
>Vancouver, B.C.
>Canada  V6T 1Z3
>
>Office phone: (604) 822-8086
>Office facsimile: (604) 822-8693
>Confidential home facsimile: (604) 272-2102
>E-mail: spelech@home.com

Hi Steve and listmates,

To your list you could add
Rsk3
Wee1
Myt1
Cdc2
Cdk2
Eg2

Cheers and best regards
Jim



-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-
James E. Ferrell, Jr.
ferrell@cmgm.stanford.edu