...And after coming back to the lab from the library, I would try the
following:
1. Prepare H-Phe-OH sodium salt (by adding an equivalent of NaOH or
Na2CO3 to Phe in water).
2. Prepare N-hydroxysuccinimide ester of a protected tyrosine
derivative (I would recommend Boc-Tyr(OtBu), but you may take cheaper
Boc-Tyr(OBzl)). Check in the literature if it can be obtained
crystalline; if not, you can use the oil obtained from the reaction of
the protected AA, HONSu and DCC.
3. Evaporate the solution of H-Phe-OH sodium salt to minimum amount
and add the solution of the active ester in DMF. Check that pH of the
aqueous extract from your reaction mixture is above 7 (by just putting
a drop of it on wet indicator paper). Leave the reaction mixture
overnight (under stirring if not homogenous).
4. Evaporate the mixture, add some EtOac and dilute (1-5%) aqueous
H2SO4 and separate the layers. Check that your product does not go to
the aqueous media.
5. If it stays in EtOAc, wash the organic phase with dilute H2SO4 and
saturated Na2SO4 (to remove the acid). If not, evaporate EtOAc and
purify the product by flash chromatography on silica gel using eluents
consisting of CHCl3 and MeOH with some HOAc (1-5%).
6. Try to obtain your product solid by
precipitations/crystallizations. If this does not work, go on with the
oil.
7. Remove the protectiong groups (by TFA if using Boc/tBu or
hydrogenation followed by TFA if using Boc/Bzl).
8. Purify the peptide by HPLC.
9. Control your synthesis by TLC all the way down.
10. Check the literature for specific protocols.
11. If you do not like all the jazz described above, order the peptide
from somebody for a few dollars more (well, it's really cheap).
Good luck
Vladimir
On 08/02/2001, Carter, Mark wrote:
> Enrico,
> Welcome to the wonderful world of PepSyn! I ask all my new entry level
> employees to read the excellent (if slightly dated) review by Fields and
> Noble in IJPPR as a starting point. It's probably more than you'll need for
> a single synthesis of a dipeptide, but you'll appreciate the references.
> Gregg B Fileds and Richard L Noble (1990) "Solid phase peptide synthesis
> utilizing 9-fluorenylmethoxycarbonyl amino acids," Int. J. Peptide Protein
> Res., Vol. 35, pp. 161-214.
> There are also a couple of good books around. They may be harder for you to
> find if you are using a decent library, but easier if you have to buy your
> own literature. One of the more recent comprehensive books is:
> Methods in Molecular Biology, Vol 35 Peptide Synthesis Protocols (1994) ed.
> M W Pennington and B M Dunn, Humana Press, Totowa, New Jersey.
> and its companion, Vol 36 Peptide Analysis Protocols, ed. B M Dunn and M W
> Pennington.
> Mark
> -----Original Message-----
> From: Enrico Millo [mailto:millo@vega.cba.unige.it]
> Sent: Wednesday, February 07, 2001 11:41 AM
> To: Recipients of ABRF List
> Subject: information request
> Dear ABRF,
>> my research field is peptide synthesis. I have to make a dipeptide
>>formed by Tyr-Phe in solution phase, can you give me some
>>information about a protocol to use or some reference about this
>>synthesis(reagent, conditions of coupling, best solvent)?
>>Thank a lot
>>Enrico Millo
Vladimir Titov
Bokiron Ltd., Moscow, Russia
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