PHOSPHORYLATION OF CYTADHERENCE-ACCESSORY PROTEINS IN MYCOPLASMA-PNEUMONIAE

Attachment to host cells of the respiratory epithelium by Mycoplasma p neumoniae is a complex, multicomponent process, requiring a number of accessory proteins in addition to adhesins directly involved in recept or binding. In this study, protein phosphorylation of the cytadherence -accessory proteins HMW1, HMW2, and HMW4 of M. pneumoniae was examined using biochemical and immunological techniques. The initial indicatio n of protein modification came from Western immunoblot analysis of the two-dimensional polyacrylamide gel electrophoresis (PAGE) profile of M. pneumoniae proteins, revealing multiple spots for both HMW1 and HMW 4 that varied in pI but not in size. M. pneumoniae cultured in the pre sence of (H3PO4)-P-32 exhibited numerous phosphorylated proteins as de tected by sodium dodecyl sulfate-PAGE and autoradiography. These inclu ded proteins corresponding to HMW1, HMW2, and HMW4 in electrophoretic mobility. The Triton X-100 partitioning characteristics of these phosp horylated proteins was identical to that described previously for HMW1 , -2, and -4. Furthermore, these protein bands were absent when a nonc ytadhering variant deficient in HMW1-5 was examined in the same manner . Finally, the availability of antiserum to HMW1 and -4 enabled us to confirm by radioimmunoprecipitation that HMW1 and HMW4 are phosphoprot eins. Phosphoamino acid analysis of acid-hydrolyzed HWM1 and HMW2 iden tified primarily phosphothreonine and, to a lesser extent, phosphoseri ne in HMW1 and predominantly phosphoserine, with a trace of phosphothr eonine, in HMW2. Neither protein contained phosphotyrosine. HMW1-HMW5 are components of a cytoskeleton-like structure in M. pneumoniae that is thought to function in cell division, changes in cell morphology, g liding motility, and the localization of adhesins in the mycoplasma me mbrane. Phosphorylation may regulate cytoskeleton dynamics involving t hese cytadherence-accessory proteins.