Figure 1
From: Engineering the Campylobacter jejuni N-glycan to create an effective chicken vaccine
The protein-based C. jejuni N-glycan vaccine.
(A) The first 150 amino acids of the C. jejuni Cj1433c polypeptide are shown. The sequence containing the GlycoTag peptide (nine repeats of [KIDLNNT]) is located between the double asterisks (**). The bacterial N-glycosylation sequon (DLNNT) is underlined. (B) Schematic diagram of the ToxC-GlycoTag-His6 expression construct: pT7, IPTG inducible promoter; pelB, pET22b-derived pectate lyase B leader sequence; toxC, truncated, non-toxic variant of the C. diphtheria toxin; His6, hexa-histidine-tag. (C) The anticipated N-linked glycosylated fusion protein is shown. (D) Expression and purification of glycosylated ToxC-GlycoTag-His6 in the presence of pACYC184 (pgl) in E. coli BL21 (DE3). (a) A 12.5% SDS-PAGE of whole cell lysates of BL21/ToxC-GlycoTag-His6 (L) and combined elution fractions after IMAC (E). Western blots with R1-4 antiserum of (a) combined elution fractions after IMAC, after (b) anion exchange chromatography and (c) after size exclusion chromatography. Molecular weight markers (in kDa) are indicated on the left.
