Date: Thu, 16 Jan 1997 12:09:53 -0600
Message-Id: <v01530505af051d10fd46@[128.252.197.88]>
To: abrfhyp@cco.caltech.edu
From: rskubish@pharmdec.wustl.edu (Richard Skubish)
Subject: PepSyn-disulfide exchange
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>Date: Sat, 13 Jul 1996 14:55:42 +0800
>Old-To: jack.moore@ualberta.ca (Jack Moore),
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>From: schooley@unr.edu (David A. Schooley)
>Subject: Re: PepSyn-disulfide exchange
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>Jack Moore wrote:
>>I am working on a synthetic peptide that seems to be exchanging disulfides and
>>would appreciate any suggestions on how to minimize this problem. The
>>sequence
>>is CYIQNCPLGGKR. Both terminals are free and the two cys residues form a
>>disulfide. After purification, oxidation and another purification I have
>>my desired product at a purity level of greater than 95%. The problem is
>>that I can't stop the peptide from forming new oxidation products. This
>>occurs after a couple of days as a lyophilized powder. It happens on both
>>the benchtop and the freeze drier.
> This arginine vasotocin precursor should be readily convertible to
>a mix containing both the parallel and antiparellel dimers under basic
>conditions in concentrated solution. Triethylamine is sufficiently basic
>to promote dimer formation. It would not surprise me that this should
>happen in the precursor with KR present, but seems very unusual in the
>peptide lacking the KR, and as salts!
> Can you isolate the impurities for MS to see whether they are
>dimers? Maybe the dimers are forming during your oxidation. The dimers
>SHOULD be removed in the purification after oxidation, but this depends on
>how it was purified (RPLC will separate them, ion exchange might not).
> For references to characerization and deliberate synthesis of
>similar dimers (of CLITNCPRG-NH2) as authentic neuropeptides, see our
>paper:
>Proux et al., BBRC, 149, 180-186 (1987)
> For reference to du Vigneaud's synthesis of similar oxytocin
>dimers, see:
>Yamashiro et al., J. Am. Chem. Soc., 90, 3857-? (1968).
>
>David
>
>David A. Schooley
>Dept. of Biochemistry
>Univ. of Nevada
>Reno, NV 89557
>schooley@med.unr.edu
>tel: (702) 784-4136; fax (702) 784-1419
>
Richard Skubish
rskubish@pharmdec.wustl.edu
314-362-0283
Washington University Medical School
Box 8103 - PNACL
660 S. Euclid
St. Louis, MO 63110