The transpeptidase activity of the essential penicillin-binding protein 2x (PBP2x) of Streptococcus pneumoniae is believed to be important for murein biosynthesis required for cell division. To study the molecular mechanism driving localization of PBP2x in live cells, we constructed a set of N-terminal GFP-PBP2x fusions under the control of a zinc-inducible promoter. The ectopic fusion protein localized at mid-cell. Cells showed no growth defects even in the absence of the genomic pbp2x, demonstrating that GFP-PBP2x is functional. Depletion of GFP-PBP2x resulted in severe morphological alterations, confirming the essentiality of PBP2x and demonstrating that PBP2x is required for cell division and not for cell elongation. A genetically or antibiotic inactivated GFP-PBP2x still localized at septal sites. Remarkably, the same was true for a GFP-PBP2x derivative containing a deletion of the central transpeptidase domain, although only in the absence of the protease/chaperone HtrA. Thus localization is independent of the catalytic transpeptidase domain but requires the C-terminal PASTA domains, identifying HtrA as targeting GFP-PBP2x derivatives. Finally, PBP2x was positioned at the septum similar to PBP1a and the PASTA domain containing StkP protein, confirming that PBP2x is a key element of the divisome complex.