EXPRESSION IN MYCOPLASMA-PNEUMONIAE OF THE RECOMBINANT GENE ENCODING THE CYTADHERENCE-ASSOCIATED PROTEIN HMW1 AND IDENTIFICATION OF HMW4 ASA PRODUCT

Mycoplasma pneumoniae is a major cause of tracheobronchitis and pneumo nia in older children and young adults. The lack of adequate tools for genetic analysis has hindered the elucidation of function and regulat ion of mycoplasma virulence determinants. We describe here the use of a transposon vector to deliver the cloned gene for the cytadherence-as sociated protein HMW1 in M. pneumoniae. A 4.95 kbp BamHI fragment enco ding all but the C-terminal end of HMW1 was cloned into a modified Tn4 001 and transformed into wild-type M. pneumoniae and into a non-cytadh ering mutant lacking HMW1-HMWB. Southern blot hybridizations confirmed insertion of the transposon and the presence of both the resident and recombinant hmw1 alleles. Analysis by Western immunoblotting revealed a truncated HMW1 (HMW1') in the transformants, the level of HMW1' bei ng dependent upon the orientation of the hmw1 gene in the transposon a nd the site of insertion. Similar expression patterns were noted in wi ld-type and mutant backgrounds. However, expression of wild-type level s of HMW1' in the mutant did not restore adherence. Finally, HMW4 and HMW1 were shown to be products of the same gene, HMW4 being a heat-mod ified derivative of HMW1.