JMSSJ On-line

JMSSJ On-line, Vol. 51 (2003) No. 1, pp. 67-71

Protein and Synthesized Peptide pI Marker Analysis by Capillary Isoelectric Focusing Separation/Mass Spectrometry Using Sonic Spray Ionization		>> Full Text PDF >> References
*Yukiko HIRABAYASHI,^a) Atsumu HIRABAYASHI,^b) and Kiyohito SHIMURA^c)** ^*a)Advanced Research Laboratory, Hitachi, Ltd. (2520 Akanuma, Hatoyama-cho, Saitama 350-0395, Japan) E-mail: hiraby@crl.hitachi.co.jp ^b)Central Research Laboratory, Hitachi, Ltd. (1-280 Higashikoigakubo, Kokubunji, Tokyo 185-8601, Japan) ^c)Faculty of Pharmaceutical Sciences, Teikyo University (Sagamiko, Kanagawa 199-0195, Japan)

Protein and synthesized-peptide pI marker analysis by using capillary isoelectric focusing separation/mass spectrometry (CIEF/MS) with a sonic spray ionization (SSI) interface, which has a buffer reservoir between the ion source and the separation capillary, has been demonstrated. Various proteins have often been used as pI markers for CIEF; however, they were unstable, so their pI could be changed from their expected ones. Therefore synthetic peptide pI markers are useful because of their higher stability and purity. However, the m/z of peptide pI markers are in the same range with those of the ions of polymers with a wide range of molecular weights due to ampholytes used for CIEF. As a result, detecting the protonated molecules of the peptide pI markers was difficult. In our work, we have optimized the scanning m/z range of a three-dimensional quadrupole (3DQ) ion-trap mass spectrometer to analyze of peptide pI markers and proteins, respectively, and to scan intensities of two ranges at the same time. Specifically, we separated the scanning range from 300 to 500 and from 700 to 1900. Using this method, carbonic anhydrase II, myoglobin, cytochrome-c, and four peptide pI markers were analyzed simultaneously by using CIEF/MS with an SSI interface, and these proteins were observed with reasonable pI values within the limit of the separation-power of this system.

Key words: Capillary isoelectric focusing separation/mass spectrometry (CIEF/MS), Sonic spray ionization, Protein, Peptide pI marker, Proteomics

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