Real-Time Monitoring of Recombinant Bacterial Proteins by Mass Spectrometry

• Published in: Biotechnology progress 2005-11, Vol.21 (6), p.1754-1758
• Main Authors: Jones, Jeffrey J., Wilkins, Charles L., Cai, Ying, Beitle, Robert R., Liyanage, Rohanna, Lay Jr, Jackson O.
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - analysis; Bacterial Proteins - genetics; Biological and medical sciences; Biotechnology; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Green Fluorescent Proteins - analysis; Green Fluorescent Proteins - genetics; Recombinant Proteins - analysis; Recombinant Proteins - genetics; Spectrometry, Fluorescence; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
• ISSN: 8756-7938; 1520-6033
• DOI: 10.1021/bp050108o

Genetically altered bacteria manipulated to express green fluorescent protein (GFP) were used in an investigation of real‐time monitoring for recombinant protein expression in cell by matrix‐assisted laser desorption/ionization (MALDI) time‐of‐flight (TOF) mass spectrometry (MS). A significant advantage to whole cell MALDI MS is its ability to analyze bacterial cultures without pretreatment other than concentration. This paper describes the simultaneous analysis of overexpressed GFP recombinant Escherichia coliJM101 by MALDI‐TOF MS and standard fluorescence measurements. Cells were harvested from liquid culture media during a 12 h GFP induced expression cycle to demonstrate the feasibility of near real‐time monitoring of induced protein expression. The results show that although MALDI MS is not as sensitive as fluorescence measurements, expression levels of the targeted protein can easily be determined. Data available only through MALDI MS measurements reveal the presence of both native GFP and GFP‐(histidine)6 proteins. Additionally, biochemical processes not yet fully understood are observed in the presence and absence of ribosomal protein constituents. Thus, the work presented here demonstrates the ability of MALDI MS to monitor and characterize in real time the expression of targeted and unexpected genetically recombinant proteins in active cell cultures.