Use of double-depleted 13C and 15N culture media for analysis of whole cell bacteria by MALDI time-of-flight and Fourier transform mass spectrometry.
In the present paper, results demonstrating the significant advantages of matrix-assisted laser desorption/ionization (MALDI) analysis of whole cell samples of bacteria grown on double isotopically-depleted (13C and 15N) media are presented. It is shown that several advantages accrue for MALDI with a 9.4 T Fourier transform mass spectrometer (FTMS). Of particular note, for analysis of whole cells, sample preparation is simple and chemical interference is reduced. Moreover, ion coalescence problems are minimized, and data-base identification of proteins facilitated. Furthermore, high resolution mass spectra obtained from such whole cells show significant improvement in apparent mass resolving power and mass measurement accuracy, whether time-of-flight or FTMS MALDI is used. As a consequence, it becomes possible to detect subtle details in the chemistry of the organism, such as the presence of both post-translationally modified and unmodified versions of the same proteins. This approach is also adaptable to direct assay of over-expressed proteins from Escherichia coli cultures and should facilitate studies aimed at the detection of medically important cellular biomarker proteins.
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, Arkansas 72701, USA.
Michael J
MJ
Stump
Jeffrey J
JJ
Jones
Richard C
RC
Fleming
Jackson O
JO
Lay
Charles L
CL
Wilkins
D001419:Bacteria / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry; D002138:Calibration; D002247:Carbon Isotopes / Q000032:analysis; D003470:Culture Media / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry; D005583:Fourier Analysis; D009587:Nitrogen Isotopes / Q000032:analysis; D012270:Ribosomes / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001426:Bacterial Proteins; D002247:Carbon Isotopes; D003470:Culture Media; D004268:DNA-Binding Proteins; D009587:Nitrogen Isotopes