Exploring Metalloproteome Remodeling in Calprotectin-Stressed Acinetobacter baumannii Using Chemoproteomics.

Area of Science:

• Microbiology and Immunology
• Proteomics and Biochemistry

Background:

• Nutritional immunity is a host defense mechanism that restricts pathogen growth by limiting access to essential transition metals.
• Calprotectin (CP) is a key host protein involved in nutritional immunity, known to scavenge zinc (Zn) and iron (Fe).
• Previous studies showed CP induces Zn- and Fe-starvation responses in *Acinetobacter baumannii*.

Purpose of the Study:

• To quantitatively assess changes in cysteine reactivity and cellular metal occupancy in metalloenzymes of *A. baumannii* under CP stress.
• To identify specific cellular processes and proteins affected by metal limitation induced by CP.

Main Methods:

• Quantitative chemoproteomics platform used to measure abundance-corrected cysteine reactivity in *A. baumannii*.
• Comparison of CP-stressed *A. baumannii* to untreated wild-type (WT) control.
• Proteomics analysis performed on a CP-stressed Δ*zigA* strain relative to a CP-stressed WT strain to investigate the role of ZigA.

Main Results:

• CP stress induced a pronounced Zn-limitation and Fe-starvation response, with reciprocal regulation of central carbon metabolism enzymes like aconitase.
• A significant decrease in metal occupancy (undermetalation) was observed across the proteome, affecting known Zn-, Fe-, and Fe-S-cluster proteins.
• Undermetalation impacted diverse cellular processes including the TCA cycle, respiration, GTP metabolism, ribosome remodeling, tRNA charging, and proteostasis.

Conclusions:

• Calprotectin induces widespread undermetalation of the metalloproteome in *Acinetobacter baumannii*.
• This widespread undermetalation leads to significant nutrient limitation and impacts numerous vital cellular functions.
• The metallochaperone ZigA does not appear to be a primary client enzyme affected by CP-induced undermetalation in this assay.