Fluorescence detection of amino acids in the postcleavage conversions for manual sequencing of a peptide

Wainaina, Moses N.; Shibata, Takayuki; Smanmoo, Chaivat; Kabashima, Tsutomu; Kai, Masahiro

doi:10.1016/j.ab.2007.12.003

Cited by 12 publications

(12 citation statements)

References 9 publications

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“…Alternatively, modified Edman degradation was utilized in order to investigate the possibility of the detection of phosphoamino acid with PITC, in which peptides were reacted with PITC and the obtained ATZ-amino acid intermediates were treated with CIA to generate fluorescent PTC-CIA derivatives [26]. However, the PTC-CIA corresponding to the phosphoamino acids could not be detected and identified (data not shown).…”

Section: Identification Of Cip-tz Amino Acidsmentioning

confidence: 99%

“…Furthermore, multiple product peaks appeared in the case of Thr(p) and Ser(p), and they contained identical peaks to the CIA derivatives of parent amino acids, possibly due to dephosphorylation. The reaction for fluorescence derivatization of ATZ-amino acids with CIA requires heating at 80˚C for 20 min [26]. This harsh reaction condition should cause the dephosphorilation and/or the degradation of phospho-PTC-CIA.…”

Section: Identification Of Cip-tz Amino Acidsmentioning

confidence: 99%

“…We have previously reported the manual sequencing method of peptide utilizing modified Edman degradation technique [26]. In this work, a small filter disk made of polypropylene was employed as a reaction vessel.…”

Section: Derivatization Conditions With Cipicmentioning

confidence: 99%

“…At the cleavage stage of the CIPIC reaction to generate CIP-TZ-amino acids, we applied a gas-phase method which was found to reduce the generation of TFA-related impurities on membranes [26]. The reaction vessel was tightly attached to a tube containing liquid TFA, and TFA vapor generated by heating the tube was allowed to pass through the membranes.…”

Section: <Fig 3>mentioning

confidence: 99%

“…Edman degradation [26], although the 2nd, 3rd and 4th residues of the pentapeptide were 5 characterized. These results clearly indicated that CIPIC affords the first example of the sequencing of phosphopeptide based on Edman degradation protocol, and the applicability of the present method for direct determination of phosphorylated amino acid residue in phosphoproteins.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

A manual sequence method of peptides and phosphopeptides using 4-(1′-cyanoisoindolyl)phenylisothiocyanate

Shibata

Wainaina

Miyoshi

et al. 2011

Journal of Chromatography A

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A method for sequence analysis and identification of phosphoamino acids in peptides based on high performance liquid chromatography (HPLC) is described. The peptides were derivatized with an Edman type reagent, 4-(1'-cyanoisoindolyl)phenylisothiocyanate (CIPIC) and subsequently cleaved to generate stable and fluorescent 4-(1'-cyanoisoindolyl)phenylthiazolinone (CIP-TZ)-amino acids. Several experimental factors that affected derivatization on membranes were examined. Under the optimized conditions, the CIP-TZ derivatives of Try(p), Thr(p) and Ser(p) were obtained and separated from their parent amino acids with baseline resolution using an isocratic elution system. Up to the 4th residue of phosphorylated pentapeptides was successfully identified, whereas phosphoamino acid residues could not be detected by the conventional procedure using phenylisothiocyanate (PITC). The results demonstrated the potential of CIPIC as a derivatization reagent for peptide sequencing and the applicability of the method for the study and identification of phosphoamino acids in peptides.

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Section: Identification Of Cip-tz Amino Acidsmentioning

confidence: 99%

Section: Identification Of Cip-tz Amino Acidsmentioning

confidence: 99%

Section: Derivatization Conditions With Cipicmentioning

confidence: 99%

Section: <Fig 3>mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

A manual sequence method of peptides and phosphopeptides using 4-(1′-cyanoisoindolyl)phenylisothiocyanate

Shibata

Wainaina

Miyoshi

et al. 2011

Journal of Chromatography A

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Synthesis of 1‐(1H‐Tetrazol‐5‐yl)‐2H‐isoindole Derivatives through Ugi Four‐Component and Silver‐Catalyzed Reactions

Gao

et al. 2014

Eur J Org Chem

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A facile and efficient one‐pot, two‐step synthesis of 1,2‐disubstituted 3‐(1H‐tetrazol‐5‐yl)‐2H‐isoindoles through Ugi four‐component reaction/AgNO3‐catalyzed reaction of 2‐alkynylbenzaldehydes, amines, isonitriles, and Me3SiN3 is described. This transformation proceeds through 5‐exo‐dig cyclization, [1,3]‐H shift, and [1,5]‐H shift to generate the products in moderate to excellent yields.

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Analysis of PTH‐Cysteine by Adsorptive Stripping Square‐Wave Voltammetry

et al. 2009

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The aim of this work is the study of cysteine by its derivative form. Cysteine derivation is done by Edman degradation and we obtain the phenylthiohydantoin of cysteine. Edman degradation is one of the most usual procedures for sequentate proteins. From this reaction, the phenylthiohydantoin amino acid derivative (PTH-amino acid) from the N-terminal one is obtained, leaving the new amino terminus for the next degradation cycle. To identify the produced PTH-derivative, a square-wave cathodic stripping voltammetry method is developed. In this work we show the results obtained for the optimization the instrumental and chemical conditions, and we can conclude that it is an electrochemically irreversible system with adsorptive reduction phemomenon. Under the best conditions we found a new method to analyze proteins with LOD of 2 mg L À1 and LOQ of 5 mg L À1 , a RSD less than 10% and a E r minor than 9%. Results of the application of this methodology to a real protein are included.

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Fluorescence detection of amino acids in the postcleavage conversions for manual sequencing of a peptide

Cited by 12 publications

References 9 publications

A manual sequence method of peptides and phosphopeptides using 4-(1′-cyanoisoindolyl)phenylisothiocyanate

A manual sequence method of peptides and phosphopeptides using 4-(1′-cyanoisoindolyl)phenylisothiocyanate

Synthesis of 1‐(1H‐Tetrazol‐5‐yl)‐2H‐isoindole Derivatives through Ugi Four‐Component and Silver‐Catalyzed Reactions

Analysis of PTH‐Cysteine by Adsorptive Stripping Square‐Wave Voltammetry

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