The effect of substrate on glycoconjugate synthesis by <i>M</i>. <i>pulmonis</i> and <i>M</i>. <i>pneumoniae</i>.

<p>Panels A and B show the relative abundance of glucose or xylose, respectively, linked to protein as determined by GC. The values represent averages of the areas under the curves from 3 replicates of gas chromatograms that were converted to μg of sugar per mg of protein. The colors of the bars representing the different substrates used to supplement the medium are shown on the right. Plus or minus standard error bars are shown and the asterisks indicate a significant difference between a sample and control.</p

LC MS/MS-CID showing hexosylation at Asn335 of the peptide STLEYTINNSQELQn₃₃₅ILKQTYEEFTK of MYPU_3200.

<p>The assigned b and y ions are shown in blue and red, respectively. Glycosylation of N, Q, T, S, and Y glycosites is absent in this spectrum as illustrated. The PEAKS peptide score (-10lgP) for this spectrum was 60. The charge state of the parental ion was <i>z</i> = 3.</p

Hexosylation of the peptide STLEYTINNSQELQNILKQTYEEFTK of MYPU_3200.

<p>Orbitrap MS showing the doubly and triply charged ions. The monoisotopic mass of the doubly charged species at 1648.8077 is consistent with the hexosylated peptide at <i>z</i> = 2 with a mass accuracy of 0.0012 Da. The 54.0175 shift for <i>z</i> = 3 between non-glycosylated and hexose forms equates to a mass shift of 162.0525 Da with a mass accuracy of 0.0003 Da. Monoisotopic values for the calculated theoretical and experimental masses of the peptide are given in bold. The images presented were obtained from an LC peak of MS scans and are expanded to show the charge states of each form.</p

LC MS/MS-CID showing hexosylation at Thr64 of the peptide Gt₆₄KDFLPIELQSLEVSK of MYPU_3230.

<p>The assigned b and y ions are shown in blue and red, respectively. Glycosylation of Q and S glycosites is absent in this spectrum as illustrated. The PEAKS peptide score (-10lgP) for this spectrum was 87. The charge state of the parental ion was <i>z</i> = 3.</p

Hexosylation of the peptide GTKDFLPIELQSLEVSK of MYPU_3230.

<p>Orbitrap MS showing the doubly and triply charged ions. The 81.0262 shift for <i>z</i> = 2 between the non-glycosylated and glycosylated peptides equates to a mass shift of 162.0524 Da, which corresponds to the addition of hexose (162.0528 Da) with a mass accuracy of 0.0004 Da. The 54.0169 shift for <i>z</i> = 3 between non-glycosylated and glycosylated forms equates to a mass shift of 162.0507 Da, which corresponds to hexosylation with a mass accuracy of 0.0021 Da. Monoisotopic values for the calculated theoretical and experimental masses of the peptide are given in bold. The images presented were obtained from an LC peak of MS scans and are expanded to show the charge states of each form.</p

LC MS/MS-CID showing hexosylation at Gln49 of the peptide ITDLLSq₄₉KEVTETQK of MYPU_3460.

<p>The assigned b and y ions are shown in blue and red, respectively. Glycosylation of Q, S, and T glycosites is absent in this spectrum as illustrated. The PEAKS peptide score (-10lgP) for this spectrum was 74. The charge state of the parental ion was <i>z</i> = 3.</p

Hexosylation of the peptide ITDLLSQKEVTETQK of MYPU_3460.

<p>Orbitrap MS showing the doubly and triply charged ions. The 81.027 shift for <i>z</i> = 2 between the non-glycosylated and glycosylated peptides equates to a mass shift of 162.054 Da, which corresponds to the addition of hexose (162.0528 Da) with a mass accuracy of 0.0012 Da. The 54.0177 shift for <i>z</i> = 3 between non-glycosylated and glycosylated forms equates to a mass shift of 162.0531 Da, which corresponds to hexosylation with a mass accuracy of 0.0003 Da. The calculated theoretical and experimental values for <i>m/z</i> are given in bold. The images presented were obtained from an LC peak of MS scans and are expanded to show the charge states of each form.</p

Glycosites identified in this study.

<p>^a Accession numbers for MYPU_3200, MYPU_3230, MYPU_3460, and MARTH_403 are CAC13493, CAC13496, CAC13519, and YP_001999972, respectively.</p><p>Glycosites identified in this study.</p

Additional file 1: Table S1. of Comparative genome analysis of Mycoplasma pneumoniae

Initial assembly characteristics. (XLSX 19 kb

Additional file 4: Table S4. of Comparative genome analysis of Mycoplasma pneumoniae

Variants in ORF6 excluding the large variation region (nucleotides 186572-187144), relative to the M129 reference strain. (XLSX 11 kb