6 replies to this topic

[anonymous]

Well, I pressume that you measured the size by SDS-PAGE. Sometimes you get charged residues that actually change the electrophoretic pattern of your protein (it happens in my protein). I guess glycosylation might as well explain it.

[anonymous]

How big is your protein?And how big it appears to be by your method?25KD difference is really too big.

[anonymous]

Hi !

I expressed cell membrane permeable TAT fusion proteins in E coliexpression is good, functionally also it is active but the size is very different from mammalian protein. Approx. 25 kDadifference in size. Can anybody suggest something about this discripancy in size.What kind of functional changes it may impose. Though, I am sure this is the same protein but the size is different.Please suggest !Thanks!

[anonymous]

Well, I presume that you measured the size by SDS-PAGE. Sometimes you get charged residues that actually change the electrophoretic pattern of your protein (it happens in my protein). I guess glycosylation might as well explain it.

[anonymous]

Hi!The size of my protein in mammalian cells is 80kD wereas in E.coliit comes out to be 110 kD . Precisely, there are 4 variants of these proteins and they all are different in size by 5-15kDA among each other in mammalian system. The same pattern is also seen in E coli when they are expressed as TAT fusion proteinsbut here in case of each variant the size is different by 25 kDA.This looks surprizing. Though one should expect the difference of 5 kD as it is TAT fusion protein, and difference should be because of additional 16 amino acids but I do not understand the reason of this much difference. Can anybody suggest me something . Please respond ASAP!Thanx!

[anonymous]

Hi!The size of my protein in mammalian cells is 80kD wereas in E.coliit comes out to be 110 kD . Precisely, there are 4 variants of these proteins and they all are different in size by 5-15kDA among each other in mammalian system. The same pattern is also seen in E coli when they are expressed as TAT fusion proteinsbut here in case of each variant the size is different by 25 kDA.This looks surprizing. Though one should expect the difference of 5 kD as it is TAT fusion protein, and difference should be because of additional 16 amino acids but I do not understand the reason of this much difference. Can anybody suggest me something . Please respond ASAP!Thanx!

[anlicin3]

I am purifying 43kd his tagged protein from nickel column. in precast gel in sds running buffer without adding much reducing sample buffer and boiling I am running the protein along with the urea( used for elution). I got pure protein at 35kd.In PBS the protein is insoluble. can any one explain?
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