The phosphodiesterase activity of the HmsP EAL domain is required for negative regulation of biofilm formation in Yersinia pestis

Abstract

In Yersinia pestis, biofilm formation is stimulated by HmsT, a GGDEF-domain containing protein that synthesizes cyclic-di-GMP (c-di-GMP), and inhibited by HmsP, an EAL-domain protein. Only the EAL-domain portion of HmsP is required to inhibit biofilm formation. The EAL domain of HmsP was purified as a 6XHis-tag fusion protein and demonstrated to have phosphodiesterase activity using bis(p-nitrophenyl) phosphate (bis-pNPP) as a substrate. This enzymatic activity was strictly manganese dependent. A critical residue (E506) of HmsP within the EAL domain, that is required for inhibition of biofilm formation, is also essential for this phosphodiesterase activity. While the proposed function of EAL-domain proteins is to linearize c-di-GMP, this is a direct demonstration of the required phosphodiesterase activity of a purified EAL-domain protein.