Identification of N-terminal acetylation of recombinant human prothymosin alpha in Escherichia coli.
Functional modification of protein through N-terminal acetylation is common in eukaryotes but rare in prokaryotes. Prothymosin alpha is an essential protein in immune stimulation and apoptosis regulation. The protein is N-terminal acetylated in eukaryotes, but similar modification has never been found in recombinant protein produced in prokaryotes. In this study, two mass components of recombinant human prothymosin alpha expressed in Escherichia coli were identified and separated by RP-HPLC. Mass spectrometry of the two components showed that one of them had a 42 Da mass increment as compared with the theoretical mass of human prothymosin alpha, which suggested a modification of acetylation. The mass of another one was equal to that of the theoretical one. Peptides mass spectrometry of the modified component showed that the 42-Da mass increment occurred in the N-terminal peptide domain, and MS/MS peptide sequencing of the N-terminal peptide found that the acetylated modification occurred at the N-terminal serine residue. So, part of the recombinant human prothymosin alpha produced by E. coli was N-terminal acetylated. This finding adds a new clue for the mechanism of acetylated modification in prokaryotes, and also suggested a new method for production of N-terminal modificated prothymosin alpha and thymosin alpha1.
Wu
Jun
J
Beijing Institute of Biotechnology, Fengtai District, Beijing 100071, China. Wujun@nic.bmi.ac.cn
Chang
Shaohong
S
Gong
Xing
X
Liu
Dianxin
D
Ma
Qingjun
Q
D000107:Acetylation; D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013947:Thymosin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
D011498:Protein Precursors; D011994:Recombinant Proteins; C040782:prothymosin alpha; D013947:Thymosin