A POSSIBLE ZINC PERIPLASMIC METALLOCHAPERONE OF THE CATION DIFFUSION FACILITATOR YIIP IN Pseudomonas aeruginosa

MIHELJ, PAULA; RAIMUNDA, DANIEL; CARRIZO, MARIA E

Virtual

Congreso; LIV Reunión Anual de la Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB); 2021

Transition metals (TM) are fundamental in the cellular physiology of all living organisms. Zinc is one of them and participates as enzyme co-factor and signaling pathways. Due to their chemical properties Zn2+ can become extremely toxic. Thus, cells require a fine tuning of the metal allocation. Zn2+ transporters assisted by metallochaperones are key players in this process, and in gram-negative bacteria, YiiP, a member of the Cation Diffusion Facilitator (CDF) family exports Zn2+ from the cytosol to the periplasm. However, there is yet no evidence of a partnering metallochaperone. Here we provide evidences that in Pseudomonas aeruginosa, YiiP/PA3963 participates in a Zn-dependent signaling pathways assisted by a metallochaperone encoded by PA3962 locus. Bioinformatic studies showed that PA3962 displays a 3D structural similarity with both CopZ, a Cu+ metallochaperone from Bacillus subtilis, and the N-terminal soluble domain of Zn2+-PIB-ATPases. A BLAST analysis shows that this protein is unique in the clade of Pseudomonadales and that a lipobox recognized by a Signal Peptidase II is conserved throughout all homolog members. Several amino acids candidates for Zn2+ coordination were identified. In order to assess the functional role of PA3962 we quantified the Zn content in cellular fractions of P. aeruginosa WT vs the insertional mutant PA3962::Tn5 strains. The data showed an accumulation of the metal in the inner membrane fraction of PA3962::Tn5, but not in other fractions. As observed for yiiP::Tn5, the strain PA3962::Tn5 has an increased sensitivity to imipenem. We plan further structure-function studies on PA3962 Zn2+, and other TMs, binding capacity. Finally, we discuss the role of YiiP and PA3962, hereafter periplasmic metallochaperone of YiiP (PmcY), in the context of Zn2+ signaling pathways in the P. aeruginosa physiology. In our working model, YiiP/PmcY supply Zn2+ to the two-component system CzcR/CzcS, which senses periplasmic Zn and represses the transcription of the imipenem-permeable porin OprD in the outer membrane.