The effect of Wag31 phosphorylation on the cells and the cell envelope fraction of wild-type and conditional mutants of Mycobacterium smegmatis studied by visible-wavelength Raman spectroscopy.
Non-surface-enhanced Raman spectroscopy using a 514.5 nm wavelength laser has been used to measure the molecular difference of conditional mutants of Mycobacterium smegmatis expressing three different alleles: wild-type wag31(Mtb), phosphoablative wag31T73A(Mtb), and phosphomimetic wag31T73E(Mtb). This study demonstrates that the phosphorylation of Wag31, a key cell-division protein, causes significant differences in the quantity of amino acids associated with peptidoglycan precursor proteins and lipid II which are observable in the Raman spectra of these cells. Raman spectra were also acquired from the isolated P60 cell envelope fraction of the cells expressing wag31T73A(Mtb) and wag31T73E(Mtb). A significant number of the molecular vibrational differences observed in the cells were also observed in the cell envelope fraction, indicating that these differences are indeed localized in the cell envelope. Principal component analyses and discriminant function analyses were conducted on these data to demonstrate the ease of spectral classification and the reproducibility of the data.
Hamasha
Khozima
K
Department of Physics & Astronomy, Wayne State University, 666 W Hancock, Detroit, MI 48201, USA.
Sahana
Moodakare Bheema
MB
Jani
Charul
C
Nyayapathy
Seeta
S
Kang
Choong-Min
CM
Rehse
Steven J
SJ
D000596:Amino Acids; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D020102:Mycobacterium smegmatis / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D013059:Spectrum Analysis, Raman; D014538:Uridine Diphosphate N-Acetylmuramic Acid / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
D000596:Amino Acids; D018797:Cell Cycle Proteins; D014538:Uridine Diphosphate N-Acetylmuramic Acid; C069249:muramyl-NAc-(pentapeptide)pyrophosphoryl-undecaprenol