Poor coupling

[yuanhenglidlut]

Hi guys!
I have a question concerning amino acid coupling. I find that the coupling of Fmoc-Arg(pbf)-OH very slow. I have tried several coupling reagents: at the beginning i used DIC/HOBt protocol. After one night's coupling, i found that the resin beads for the Kaiser Test were blue and solution dark blue. Then i switched to PyBop/HOBt/DIEA, after about 6h's coupling it still didn't work. Then i used TBTU/HOBt/DIEA, after a few hours' coupling, there didn't seem to be any change. At last, I used HATU/HOBt/DIEA protocol. However, after about 16h's coupling, the beads were still blue and solution darkk blue.

I don't know what to do to make the coupling of Fmoc-Arg(pbf)-OH go completition. Can anyone here give me some suggestions? I almost cried the time i saw the result of this coupling. I am now in great puzzle.

[dcc01]

can you show me the peptide sequence? The sequence has great effect on it's synthesis

[yuanhenglidlut]

Originally Posted by dcc01

can you show me the peptide sequence? The sequence has great effect on it's synthesis

RISSSSGLGCKVLRRH

The Fmoc-Arg(pbf)-OH I mentioned was at the N-terminal of this sequence. I don't know if we can predict the poor coupling of the sequence. Still puzzled!

[AplaGen]

Dear yuanhenglidlut,

Some questions:
(1) Why did you use HOBt in the Pybop-approach? I do not see a chemical reason to use HOBt - the only effect it might have would be to adjust the pH which could be done cheaper by using less DIEA.
(2) Using HOBt in combination with other coupling reagents (many people do that) makes even less sense. In case of HATU you should use HOAt.
(3) Did you cleave a sample from the resin and analyze it? Sometimes Kaiser does not show the truth. In case it is a difficult coupling: did you try different Arg-derivatives? Different solvents?

Good luck,

AplaGen

[ask03]

Originally Posted by dcc01

can you show me the peptide sequence? The sequence has great effect on it's synthesis

hi dcc01,

u mentioned that sequence of a peptide plays a great role in peptide synthesis. do you know of any bioinformatics tool which might predict percentage success of the synthesis? if not how can u tell??

thanks,

ask03

[musgun]

Originally Posted by ask03

hi dcc01,

u mentioned that sequence of a peptide plays a great role in peptide synthesis. do you know of any bioinformatics tool which might predict percentage success of the synthesis? if not how can u tell??

thanks,

ask03

I would gladly pay a good deal of money for that tool Unfortunatelly there is none that I know. Study chemistry

[ask03]

yea...sad but true...i did a lot of searchin but wanted to be sure that i havent missed out anything...thanks for ur reply

[glbiochem]

I felt your difficulty were not the Fmoc-Arg(pbf)-OH, but previous sequence.
After coupled ISSSSGLG, I believe the latter part would be hard to couple.

My suggestion is wash the resin with DMF and repeat the coupling twice or 3 times.
Block with Acetic anhydride.

[glbiochem]

Peptide companion is a free software to predict the peptide difficulty.

[ask03]

Originally Posted by glbiochem

Peptide companion is a free software to predict the peptide difficulty.

hey thanks so much glbiochem. I have come across this before and tried the demo version but not much in detail. have you used it before? how good is it?