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KMID : 0380219930260020145
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Journal of Biochemistry and Molecular Biology
1993 Volume.26 No. 2 p.145 ~ p.150
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Selective Isolation of C-terminal Peptide from Tryptic Digests of rhEGF by Immobilized Anhydrotrypsin for C-terminal Region Amino Acid Sequence Determination
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Soonha Kim
Kyu Don Kim/Kyedudk Kim and Joon Kim
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Abstract
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We have studied a novel method for selective isolation. of the C-terminal fragment from tryptic digests of recombinant human Epidermal Growth Factor (rhEGF) for C-terminal amino acid sequence determination. The peptides from the tryptic digests of rhEGF were fractionated by affinity chromatography on a column packed with immobilized anhydrotrypsin. The Arg and Lys free C-terminal peptide of rhEGF was recovered from a flow-through fraction, while the remainders were adsorbed on the column. This flow-through fraction was subjected to reverse phase high-performance liquid chromatography (RP-HPLC) in order to identify the C-terminal peptide peak in the total peptide map. Using this method, we have successfully isolated the C-terminal peptide from tryptic digests of rhEGF. The molecular weight and amino acid sequence of this fragment was identified as 633 and Trp-Trp-Glu-Leu using mass spectrometer and Protein/Peptide Sequencer. This result matches with the reported C-terminal peptide sequence (residue 49-52) of rhEGF.
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