pH and Cation-induced Thermodynamic Stability of Human Hyaluronan Binding Protein 1 Regulates Its Hyaluronan Affinity

Abstract

The diverse biological roles of hyaluronan, a complex polysaccharide in vertebrates, are now established beyond any doubt. These include acting as a vital structural component of connective tissues, the formation of loose hydrated matrices that allow cells to divide and migrate during development, and in intracellular signaling (1-5). Such variable activities may result from its interaction with a family of proteins known as hyaladherin (2).Hyaluronan-binding protein 1 (HABP1), 1 one of the members of hyaladherin family was identified (6) and its role in different cellular processes like cell adhesion and tumor invasion, sperm maturation, and motility (7-11) are under intensive investigation in our laboratories. Molecular cloning of human HABP1 revealed its multifunctional nature as its sequence was found to be identical with p32, a protein copurified with splicing factor SF2 and with the receptor of the globular head of complement factor C1q (gC1qR). It is represented as synonyms of p32/C1QBP (accession number NP_001203) in human chromosome 17. This molecule has generated a considerable interest in the last few years largely because of its multifarious functions as well as its localization in various subcellular compartments including cell surface in different cell types (8 -17). However, only one transcript of HABP1 was detected from different types of tissues suggesting that no other isoform(s) of HABP1 exist in different cell types (16).Studies on the crystal structure of p32/HABP1 revealed that it exists as a homotrimer, and each protomer consists of seven consecutive twisted anti-parallel ␤-sheets flanked by one NH 2 -terminal and two COOH-terminal ␣-helices and that the terminal ␣-helices have extensive intra-as well as intermolecular contacts. It has been postulated that these terminal helices are critical for maintaining the trimeric assembly and proteinprotein interactions (18). In a previous report, we have shown that HABP1 exists predominantly as a non-covalently linked trimer near physiological conditions and as a hexamer (dimer of trimers) in the oxidative environment (19). Being acidic in nature and having highly polar amino acid residues distributed asymmetrically on the surface, the electrostatic interactions in HABP1 are expected to have a role in dictating its folding topology and tertiary interactions. However, the relative role of the contribution of electrostatic interactions to protein stability, compared with that of hydrophobic interactions, has been the subject of long standing query (20). Part of the difficulty, in discriminating the contributions from electrostatic effect, stems from the fact that there are several different ways in which they contribute to the net stability of the native conformation. In addition, both attractive and repulsive electrostatic interactions are possible. In general, models to account for co-solute effects on protein stability may be classified in a number of ways; two major classifications are those in which the co-solute directly interacts with the prot...

Cleavage of hyaluronan is impaired in aged dermal wounds

“…It is unlikely this reflects lack of sensitivity given the large detection range of our standard curve, but even small changes in enzyme international units could reflect important biologic changes that are not reflected in statistical measures extrapolated from measures of optical density. Nonetheless these data suggest additional age-related changes that determine HA size in the wound dermis, such as circulating endogenous inhibitors of HYAL activity and/or HA binding proteins that protect HA against cleavage by HYAL (Jha et al, 2004; Evanko et al, 2007; Girish et al, 2009). The view that age-related deficits are multifactorial, with small decrements in many stages, is a recurring theme in studies of aging and tissue repair (Sadoun and Reed 2003; Gosain and DiPietro, 2004).…”

Hyaluronan as an Immune Regulator in Human Diseases

“…Thus, structural changes in HABP1 mediated by changes in the ionic environment are responsible for recognizing different ligands (173). The more stable conformation of HABP1 was attained at higher ionic strength or at acidic pH showed maximum affinity toward HA (171). Therefore, cellular HA-HABP1 interaction can be regulated by pH and ionic strength.…”

Changing Folding and Binding Stability in a Viral Coat Protein: A Comparison between Substitutions Accessible through Mutation and Those Fixed by Natural Selection

“…Burying polar residues contributes to folding stability since the intramolecular hydrogen bonding and van der Waals interactions of polar groups in folded proteins are more favorable than similar interactions with water in unfolded proteins [21] , [22] . Changes in ion concentration or pH also alters the thermodynamic stability [23] , [24] .…”

“…Bloom and collaborators presented a thermodynamic framework to predict the probability that a protein retains its structure after one or more random amino acid substitutions, and highly simplified models of proteins were used to support their prediction that the substitutions tend to be destabilizing [4] , [7] , [8] , [15] , [22] , [23] . A study by Tawfik and collaborators showed that about 70% of random substitutions of globular proteins are destabilizing ( ΔΔG >0 kcal/mol), and that about 20% are highly destabilizing ( ΔΔG >2 kcal/mol) [15] , [17] , [24] , [25] . In another study they found that substitutions associated with new enzymatic functions are mostly destabilizing [1] – [3] , [5] – [7] , [17] , [19] , [26] , [27] .…”

“…In this study, we determined how amino acid substitutions, accessible through a single mutation within a codon, change protein folding stability and protein-protein binding stability in a bacteriophage virus system. FoldX was used to estimate the changes in folding stability ( ΔΔG fold ) and binding stability ( ΔΔG bind ) for the coat protein F in the bacteriophage virus φX174 [7] , [8] , [12] , [15] , [24] , [25] . Folding and binding stabilities were calculated for all accessible substitutions for each amino acid residue in the major capsid protein (F).…”