Journal: Nature Communications
Article Title: Glycine, serine and threonine metabolism confounds efficacy of complement-mediated killing
doi: 10.1038/s41467-019-11129-5
Figure Lengend Snippet: Glycine-dependent mechanism for HtrE, NfrA, YhcD expression. a Relative ATP abundance of E. coli K12 in the presence or absence of 100 mM glycine for the indicated length of time ( n = 3). b Percent survival of E. coli K12 in the presence or absence of 5 mM ATP, 100 mM glycine or/and 100 µL serum ( n = 3). c Relative cAMP abundance of E. coli K12 in the presence or absence of 100 mM glycine or/and 100 µL serum ( n = 3). d Percent survival of the indicated mutants in the presence of 100 mM glycine, 100 µL serum or both ( n = 3). e Western blot of CRP in E. coli K12 exposed to 0–5 mM ATP or 0–4 mM AMP. f Effect of 100 mM glycine, 100 µL serum or both on CRP expression. g Western blot of HtrE, NfrA, YhcD in these mutants. h qRT-PCR of htrE , nfrA , yhcD in these mutants ( n = 3). i Flow cytometry quantification of anti-C9 neoantigen on outer membrane surface of the indicated mutants ( n = 3). j Percent survival of the indicated mutants in the presence of 100 mM glycine, 100 µL serum or both ( n = 3). k Western blot of HtrE, NfrA, YhcD of E. coli K12 in the presence of 100 mM glycine and the indicated amount of serum. l , m Western blot of HtrE, NfrA, YhcD of E. coli K12 ( l ) and Y17 ( m ) in the presence of 100 µL serum and the indicated concentrations of glycine. n Flow cytometry quantification of anti-C3b and anti-C9 neoantigen on E. coli K12 and Y17 in the presence or absence of 100 µL serum or 100 mM glycine or both. o , MST for the interaction of complement C3 with HtrE, NfrA, YhcD ( n = 3). i , n 50,000 cells were record with forwarding versus side scatter and were gated before data acquisition. Results in ( a – d , h – j , n , o ) are displayed as mean ± SEM, and significant differences are identified (* p < 0.05, ** p < 0.01) as determined by two-tailed Student’s t test. See also Supplementary Figs. –
Article Snippet: Fluorescein isothiocyanate (FITC) conjugated monoclonal antibody to human complement factor C3b/iC3b (Cat. HM2286, clone 3E7) or a C9 neoantigen of the terminal complement complex (TCC) (Cat.HM2167F, clone aE11) were from Hycult Biotech Inc., PA, USA.
Techniques: Expressing, Western Blot, Quantitative RT-PCR, Flow Cytometry, Two Tailed Test
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