Truncated sequence

Starting the synthesis from the nonreducing terminus, that is, using a polymer-bound donor, presents a similar problem of truncated sequences. Donors are... 

The first and the second approaches have provided some positive results but, unfortunately, they depend upon the structure of the protein. Typical examples of sequence modifications to improve stability and pharmacokinetics are the preparation of humanized antibodies, where part of the mouse sequence is substituted by the human form, and the granulocyte colony-stimulating factor muteins, where up to seven amino acids are substituted. Examples of truncated sequence proteins with improved characteristic are the 7-36 analogues of glucagon-like peptides or the 1-29 sequence growth hormone-releasing factor [1, 2]. 

The above-observed differences in the chemical reactivity of the attached functional groups in the linear polymeric supports hinder the quantitative coupling under normal conditions, even though the formation of truncated sequences due to sterically in-... 

Fluorous reverse phase silica gel (FRPSG) has been used in the purification of synthetic DNA fragments.In solid phase DNA synthesis, truncated sequences are often separated from the desired product after deprotection using HPLC or electrophoresis. In order to perform, parallel syntheses and separations of nucleotides the trityl-on purification procedure was developed, in which a lipophilic support material is used to separate the desired and undesired product, followed by deprotection. If the protecting group is labelled with a fluorous group, fiuorous-fiuorous interactions between the FRPSG and the protected nucleotide can be used to aid separation of the aqueous mixture. 

Obtaining quantitative yields in amino acid coupling steps and in the repetitive A -amino group deprotection is the main problem in stepwise solid-phase peptide synthesis (SPPS). Since purification is only possible in the last step accumulation of deleted peptides and truncated sequences occurs. Therefore these undesired compounds are present in the crude mixture after final cleavage from the solid support are usually closely related to the target peptide in their physical properties, resulting in difficult purification steps. 

The MSHOT3 (magic-sandwich higher-order truncation) sequence comprises three magic-sandwich units phase shifted by 120° with respect to one another. 

Core sequences, truncated sequences of a target peptide on the polymeric support caused by incomplete conversion in SPPS. 

Prolactin-releasing peptide (PrRP), hPrRP 31 SRTHRHSMEI RTPDIQPAWY AS RGIRPVGR Fa, a 31-neuropeptide (and the N-terminally truncated sequence... 

Truncated sequences, core sequences, incompletely synthesized peptides. Imperfect conversion during acylation or deprotection of the temporary protecting group in solid-phase synthesis may lead to mismatch sequences that lack some amino acids and truncated sequences (core sequences). They occur, when the accessibility or reactivity of the peptide bound to the solid phase is insufficient difficult sequences). Truncated sequences may be classically identified and quantified by a modified Edman degradation, also called preview analysis. Alternatively, mass spectrometry provides efficient tools for the identification of truncated sequences. 

Peptide synthesis is quite regularly accompanied by undesired side reactions, for instance by the alkylation of side chains during deprotection. In addition to such general problems for which at least partial remedies have been found, also a special problem is encountered in solid phase syntheses the formation of deletion peptides and truncated sequences . Both incomplete acylation and incomplete deprotection result in peptide chains from which one or more amino acid residues are missing. A second kind of deletion, the loss of the C-terminal dipeptide sequence, is caused by diketopiperazine formation followed by acylation of the resulting hydroxymethyl polymer (Fig. 8). Premature chain termination (truncation) takes place if acetic acid or trifluoroacetic acid are not completely removed after deprotection and then co-activated in the following coupling step. 

Though quantitative conversions on solid phase are hard to reach, besides the imperative to block truncated sequences there is another excellent chance to limit the theoretical microheterogeneity of the end product of a Merrifield synthesis. As shown above, the distribution and quantity of contaminations increase exponentially with the number of stages of an incremental synthesis, whereas the possibility of distinguishing between the main and by-products decreases in the same exponential proportion. 

Reprocess truncated sequences GPMAW Output text files containing in silico fragment ions of each sequence... 

Figure 1. Purification of rat CpnIO (101 residues) using Fmoc probe 2. (A) Analytical RP-HPLC (C4 medium) of crude underivatized rat CpnIO. (B) Addition of lipophilic probe 2 increases the retention time of the protein (labelled 2) thus facilitating purification from underivatized truncated sequences (labelled 1). (C) Purified protein derivatized with 2. (D Purified protein after treatment with 5% aqueous TEA to remove 2. (E) ESI-MS of purified rat CpnIO. (R Deconvoluted mass spectrum for purified rat CpnIO. The calculated mass of target product is 10770.57 Da (average). The found mass is 10771.0. (Q) RP-HPLC (C4 medium, gradient TFA-water into 100% TFA-AcCN, 60 min) of purified rat CpnIO. The insert shows the expanded peak. (H) CZE of purified rat CpnIO. The concentration of the major peak (protein in its native, heptameric state) is 84%. Separate size-exclusion chromatography experiments showed that the majority of the flanking peaks correspond to protein with correct sequence but having an aggregation state different from the major peak.

Diketopiperazine formation. Results in loss of C-terminal dipeptide from resin. May also result in formation of truncated sequences missing C-terminal dipeptide... 

The SPS overcomes solubility problems in the course of the synthesis, and it stands out for the simplicity and rapidity of the experimental steps as well as its potential to automatize the physical manipulations. Its major flaw is the accumulation of resin-bound impurities such as failure and truncated sequences because of incomplete coupling reactions In recent years, the SPS received new impulse by... 

Formation of error-peptides Weygand and Obermeier (1968) and Bayer et aL (1970a) have divided the error-peptides (peptides other than those desired) into two classes (a) truncated sequences and (b) failure sequences. When a peptide chain on the resin fails to grow beyond a certain length, it is termed a truncated sequence. In a failure sequence, one or more amino acids may be missing from the desired peptide sequences. 

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