Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy

Chan JW, Winhold H, Corzett MH, Ulloa JM, Cosman M, Balhorn R, Huser T (2007)
Cytometry Part A 71A(7): 468-474.

Zeitschriftenaufsatz | Veröffentlicht | Englisch
 
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Abstract / Bemerkung
Background: Laser tweezers Raman spectroscopy (LTRS) is a novel, nondestructive, and label-free method that can be used to quantitatively measure changes in cellular activity in single living cells. Here, we demonstrate its use to monitor changes in a population of E coli cells that occur during overexpression of a protein, the extracellular domain of myelin oligodendrocyte glycoprotein [MOG(1-120)]. Methods: Raman spectra were acquired from individual E coli cells suspended in solution and trapped by a single tightly focused laser beam. Overexpression of MOG(I-120) in transformed E coli Rosetta-Gami (DE3)pLysS cells was induced by addition of isopropyl thiogalactoside (IPTG). Changes in the peak intensities of the Raman spectra from a population of cells were monitored and analyzed over a total duration of 3 h. Data were also collected for concentrated purified MOG(1-120) protein in solution, and the spectra compared with that obtained for the MOG(I-120) expressing cells. Results: Raman spectra of individual, living E coli cells exhibit signatures due to DNA and protein molecular vibrations. Characteristic Raman markers associated with protein vibrations, such as 1,257, 1,340, 1,453, and 1,660 cm(-1), are shown to increase as a function of time following the addition of IPTG. Comparison of these spectra and the spectra of purified MOG protein indicates that the changes are predominantly due to the induction of MOG protein expression. Protein expression was found to occur mostly within the second hour, with a 470% increase relative to the protein expressed in the first hour. A 230% relative increase between the second and third hour indicates that protein expression begins to level off within the third hour. Conclusion: It is demonstrated that LTRS has sufficient sensitivity for real-time, nondestructive, and quantitative monitoring of biological processes, such as protein expression, in single living cells. Such capabilities, which are not: currently available in flow cytometry, open up new possibilities for analyzing cellular processes occurring in single microbial and eukaryotic cells, Published 2007 Wiley-Liss, Inc.
Stichworte
single cells; E. coli; glycoprotein; expression; protein; laser trapping; raman spectroscopy; optical tweezers
Erscheinungsjahr
2007
Zeitschriftentitel
Cytometry Part A
Band
71A
Ausgabe
7
Seite(n)
468-474
ISSN
1552-4922
eISSN
1552-4930
Page URI
https://pub.uni-bielefeld.de/record/2330530

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Chan JW, Winhold H, Corzett MH, et al. Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy. Cytometry Part A. 2007;71A(7):468-474.
Chan, J. W., Winhold, H., Corzett, M. H., Ulloa, J. M., Cosman, M., Balhorn, R., & Huser, T. (2007). Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy. Cytometry Part A, 71A(7), 468-474. doi:10.1002/cyto.a.20407
Chan, J. W., Winhold, H., Corzett, M. H., Ulloa, J. M., Cosman, M., Balhorn, R., and Huser, T. (2007). Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy. Cytometry Part A 71A, 468-474.
Chan, J.W., et al., 2007. Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy. Cytometry Part A, 71A(7), p 468-474.
J.W. Chan, et al., “Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy”, Cytometry Part A, vol. 71A, 2007, pp. 468-474.
Chan, J.W., Winhold, H., Corzett, M.H., Ulloa, J.M., Cosman, M., Balhorn, R., Huser, T.: Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy. Cytometry Part A. 71A, 468-474 (2007).
Chan, James W., Winhold, Heiko, Corzett, Michele H., Ulloa, Joshua M., Cosman, Monique, Balhorn, Rod, and Huser, Thomas. “Monitoring dynamic protein expression in living E-coli. Bacterial Celts by laser tweezers raman spectroscopy”. Cytometry Part A 71A.7 (2007): 468-474.

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