Identification of two penicillin-binding multienzyme complexes in Haemophilus influenzae

Dansyl-labeled penicillin, reversed-phase chromatography, and peptide mappi ng have been used to detect, separate, and study penicillin-binding protein s (PBPs) and PBP multienzyme complexes of H. influenzae. The cross-linking of proteins in the multienzyme complex was accomplished with the aid of cya nogen, a salt-bridge specific cross-linking agent. The chromatographic prof ile of the PBPs clearly showed a dramatic change in the number and identity of peaks after treatment of the bacterial cells with cyanogen. The disappe arance of all seven peaks corresponding to the PBPs was accompanied by the emergence of two new peaks with molecular weights between 400 kDa and 600 k Da. The results hint at the existence of two penicillin-binding multienzyme complexes, each containing subunits that interact via salt-bridges. Chroma tographic active site peptide mapping of PBPs and PBP complexes was used to determine the identity of PBPs involved in each complex. It is postulated that one multienzyme complex containing PBP 2 may be involved in cell elong ation while the other complex containing PBP 3 may be responsible for cell division. (C) 1999 Academic Press.