Fig. 11.

UMPK is an A. phagocytophilum surface protein.

A. UMPK is a 244-amino-acid protein with two predicted TM domains at residues 101–122 and 132–154. Antiserum was raised against a peptide that corresponds to amino acids 182–195, which is predicted to be surface exposed.

B. Predicted outer membrane (OM) topology of UMPK. OUT, extracellular. IN, interior to the OM.

C. UMPK localizes within the ApV. A. phagocytophilum-infected RF/6A cells, and human neutrophils were screened with antibodies against UMPK and APH0032 and visualized by confocal microscopy. The insets denoted by solid boxes are enlarged versions of the vacuoles demarcated by hatched-line boxes. Host cell nuclei and bacterial DNA were stained with DAPI (blue).

D. Proteinase K removes a surface-exposed UMPK epitope. Intact A. phagocytophilum organisms were incubated with proteinase K (+) or vehicle control (−) followed by washing to remove cleaved proteins. The bacteria were lysed and analysed by Western blot (WB) using antibodies against UMPK residues 182–195, a known-surface exposed epitope of the A. phagocytophilum surface protein, Asp55, and APH0032, which is not a surface protein. Results presented are representative of two experiments with similar results.