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Protein Size


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3 replies to this topic

#1 ABJF

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Posted 10 January 2010 - 05:38 PM

Hi,
I found that my proteins which contain a S-S bond shown larger band size in reduced gel (128kDa) than non-reduced gel (109kDa), although there are more bands appeared in reduced gel. I repeated a few times but still got the same pattern. Shouldn't it show smaller band size in reduced gel? Any opinion or explanation for these results? Appreciate any help.

Edited by ABJF, 10 January 2010 - 06:20 PM.


#2 HomeBrew

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Posted 10 January 2010 - 08:41 PM

A protein with disulfide bonds intact will be more compact, and be able to navigate the pores of the gel matrix more quickly, thus appearing to be of smaller molecular weight.

#3 ABJF

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Posted 10 January 2010 - 09:22 PM

Thanks for the reply. But I added SDS sample buffer and boiled the proteins at 95C for 5 mins, the proteins should be linearized already right? Btw, I used tris-glycine gel 8-16%.
And in reduced gel, I got three major bands: ~128 kda
~100 kDa
~20-30 kDa
Whereas in non-reduced gel, I only get one major band ~109kDa.

#4 Prep!

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Posted 11 January 2010 - 05:09 AM

wat home brew says still holds true... sds wont denature your protein per say viz.. disulfide bonds are still intact!!!
the band near 100 kDa might be your main band were sum molecules have not been reduced so may be the conc of Bme or dtt is less... more sample details might help!!!

Edited by Pradeep Iyer, 11 January 2010 - 05:11 AM.

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