Membrane penetration of the FYVE domain is modulated by pH

Abstract

The FYVE domain (Fab1, YOTB, Vac1, EEA1) binds phosphatidylinositol 3‐phosphate [PtdIns(3)P] in membranes of early endosomes and penetrates bilayers. Here, we investigate the membrane anchoring mechanism by NMR, liposome binding, monolayer surface tension, mutagenesis, and in‐vivo localization experiment. Our results show that the extent of the FYVE domain insertion into PtdIns(3)P‐enriched membranes is substantially enhanced by acidic conditions and contacts with anionic lipids. The EEA1 FYVE domain associates with POPC/POPE/PtdIns(3)P vesicles with a Kd of 49 nM at pH 6.0, however binding is ~~24 fold weaker at pH 8.0. The decrease of the binding affinity is primarily due to much faster dissociation of the FYVE domain from membranes in basic media. The membrane penetration is modulated by the tandem His residues in the RRHHCRXCG signature motif, mutations of which abolish the pH‐sensitivity. The increased membrane association in vitro and in vivo by the Hrs, RUFY1, Vps27p and WDFY1 FYVE domains in a low pH environment demonstrates that pH‐dependency is a general function of the FYVE protein family.Supported by NIH grants GM071424 and CA113472 (TGK).