Serendipitous Discovery and X-Ray Structure of a Human Phosphate Binding Apolipoprotein

Abstract

We report the serendipitous discovery of a human plasma phosphate binding protein (HPBP). This 38 kDa protein is copurified with the enzyme paraoxonase. Its X-ray structure is similar to the prokaryotic phosphate solute binding proteins (SBPs) associated with ATP binding cassette transmembrane transporters, though phosphate-SBPs have never been characterized or predicted from nucleic acid databases in eukaryotes. However, HPBP belongs to the family of ubiquitous eukaryotic proteins named DING, meaning that phosphate-SBPs are also widespread in eukaryotes. The systematic absence of complete genes for eukaryotic phosphate-SBP from databases is intriguing, but the astonishing 90% sequence conservation between genes belonging to evolutionary distant species suggests that the corresponding proteins play an important function. HPBP is the only known transporter capable of binding phosphate ions in human plasma and may become a new predictor of or a potential therapeutic agent for phosphate-related diseases such as atherosclerosis.

Crystallization, diffraction data collection and preliminary crystallographic analysis of DING protein fromPseudomonas fluorescens

“…Van Dorsselaer & E. Chabriere, in preparation). The crystallographic structure of HPBP complexed with a phosphate ion confirmed that the DING-protein family is related to PstS proteins and adopts the characteristic 'Venus flytrap' motif (Morales et al, 2006). Interestingly, some Pseudomonas species, but to our knowledge no other bacteria, have been shown to express DING proteins that are more homologous to eukaryotic DINGs (Berna et al, 2007).…”

“…A 38 kDa DING protein was isolated from breast carcinoma cells by genisteinagarose affinity chromatography (Belenky et al, 2003). Human phosphate-binding protein (HPBP), the first DING protein to also be identified as a phosphate-binding protein, was purified from a human plasma lipoprotein fraction (Morales et al, 2006). The first plant DING was identified in tobacco by virtue of its strong binding to an Arabidopsis thaliana germin-like protein (Berna et al, 2002).…”

Structure–function relationships in a bacterial DING protein

“…The pstS and the HPBP structures fit a model known as the ''Venus flytrap'', in which two globular domains hinge together to form the phosphate-binding site, with eight conserved residues Hbonding to phosphate [5,7,8].…”

Proteomics-based approach for identification and purification of human phosphate binding apolipoprotein from amniotic fluid

“…HPBP is a novel member of the DING family of proteins, which are likely ubiquitous in eukaryotes. In humans, DING protein had been identified from synovial fluid of rheumatoid arthritis (Hain et al, 1996;Blass et al, 1999), breast cells (Belenky et al, 2003) and kidney epithelial cells (Kumar et al, 2004), but the first structure was deduced from the HPBP concomitantly purified with PON1 from human serum (Morales et al, 2006). The structure clearly showed a relationship between HPBP and ABC (ATP binding cassette transmembrane) transporters in prokaryotes.…”

“…HPBP was first purified by chance along with PON1. HPBP is a member of the DING family, which suggests its distribution in a wide number of eukaryotes (Morales et al, 2006(Morales et al, , 2007. The DING protein family seems to have emerged by consensus for ubiquitous but genetically elusive proteins, usually with a molecular weight of around 40 kDa, of which the N-terminal sequence is DINGGG- (Adams et al, 2002).…”