Serine phosphorylation regulates the P-type potassium pump KdpFABC

Area of Science:

• Microbiology
• Biochemistry
• Molecular Biology

Background:

• The KdpFABC pump is crucial for bacterial survival in potassium-deficient environments.
• Transcriptional regulation of kdpFABC expression is understood, but down-regulation mechanisms are not.

Purpose of the Study:

• To elucidate the mechanism of KdpFABC pump inhibition upon restoration of potassium levels.
• To identify the regulatory elements controlling potassium homeostasis in bacteria.

Main Methods:

• Investigated KdpFABC activity in K+-rich environments.
• Utilized site-directed mutagenesis (Ser162Ala, Ser162Asp) on KdpB.
• Performed in vitro phosphatase assays.
• Analyzed the P-type pump catalytic cycle, including ATP hydrolysis and intermediate formation (E1~P).

Main Results:

• KdpFABC is inhibited in K+-rich environments via phosphorylation of Ser162 on KdpB.
• Serine phosphorylation is reversible by serine phosphatase.
• Ser162Ala mutation leads to constitutive pump activity.
• Ser162Asp mutation inactivates the pump.
• Phosphorylation abolishes K+-dependent ATP hydrolysis and halts the catalytic cycle at the E1~P intermediate.

Conclusions:

• A novel regulatory mechanism involving serine phosphorylation of KdpB down-regulates the KdpFABC pump.
• This phosphorylation event uniquely impacts P-type pump function by altering K+-dependence and blocking the catalytic cycle.
• This finding enhances understanding of bacterial potassium homeostasis and osmotic potential regulation.