Specific volume and compressibility of human serum albumin–polyanion complexes Tibor Hianik, a, * Slavomı ´ra Ponikova ´, a Jaroslava Ba ´gelÕova ´ b and Maria ´n Antalı ´k b a Faculty of Mathematics, Physics and Computer Sciences, Comenius University, 842 48 Bratislava, Slovakia b Department of Biophysics, Institute of Experimental Physics, Slovak Academy of Sciences, 043 53 Kos ˇice, Slovakia Received 22 February 2005; revised 3 October 2005; accepted 5 October 2005 Available online 21 October 2005 Abstract—The ultrasound velocimetry, densitometry, and differential scanning calorimetry have been used to study the formation of the complexes between human serum albumin (HSA) and polyanions heparin (HEP) and/or dextran sulfate (DS). The values of the ultrasound velocity and specific volume allowed us to determine the specific adiabatic compressibility, u K /b 0 , which reflects the degree of volume compressibility of the complexes. We showed that in the presence of HEP and DS the adiabatic compressibility of HSA decreases with increasing concentration of polyanions. HEP more strongly interacts with HSA than DS. pH of electrolyte in the range 4.7–8.5 weakly affects the adiabatic compressibility. Changes of compressibility of HSA can be caused by increase of the hydration due to the formation of the HSA–polyanion complexes and due to partial unfolding of HSA. The HSA–polyanion interaction resulted in decrease of phase transition temperature of the protein. This evidences about protein destabilization in the presence of polyanions. Ó 2005 Elsevier Ltd. All rights reserved. The study of the mechanisms of interaction of poly- anions with proteins has great significance for molecular biology and medicine. For example, the protein–DNA complexes are responsible for transcription of DNA, gene expression as well as for formation of specific struc- tures of chromosomes and viruses. 1,2 On the other hand, several polyanions, such as, e.g., dextran sulfate (DS), heparin (HEP), and pentosan polysulfate reveal certain therapeutic action against development of transmissible spongiform encephalopaties (TSE). It has been shown that these polyanions bind to the prion proteins and pre- vent their accumulation in animals and cells. 3 Recent study revealed that DS blocked the synthesis of prote- ase-resistant prion (PrPres). 4 Polyanions could also inhibit the entry of HIV-1 into the cells. 5 They play a substantial role in protein–protein interactions, protein folding and stabilization 6 as well as in cell–cell commu- nication 7 . Polyanions are important in the functioning of human fluid-phase complement regulators 8 and may be used also in drug delivery. 9 Protein–polyanion inter- actions are in most cases not specific. 6 On the other hand, well over a hundred so-called heparin-binding proteins have been identified. 10 However, ability of these proteins to bind other polyanions (DNA, actin, tubulin, etc.) suggests that these interactions, although of a high affinity, are not so specific. In certain cases, however, the specific binding sites exist. 6 For example, antithrombin III has specific binding site to heparin. 10 High affinity of polyanions to the proteins is attributed to the pres- ence of positively charged binding regions 11 at the pro- tein surface and depends on electrolyte pH and ionic strength. 12 The interaction between strong polyanion— heparin and bovine serum albumin (BSA) has been stud- ied by dynamic light scattering, capillary electrophore- sis, and turbidimetry. 12,13 The interaction of BSA with heparin was observed even at pH well above the isoelec- tric point of BSA (I e  4.9). 14 This evidences about exis- tence of positively charged domains at the protein surface, that is at physiological pH in general negatively charged. 14 Existence of positively charged domains has been confirmed also by computer visualization of the protein surface. 13 According to Hattori et al. 13 heparin is bound to BSA at heparin binding site. Strong interac- tion of heparin to the other protein—ferricytochrome c has been observed by differential scanning calorimetry. At low ionic strength heparin induced an important shift of the transition temperature T m from 84.1 to 59.8 °C. Bioorganic & Medicinal Chemistry Letters 16 (2006) 274–279 0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2005.10.010 Keywords: Human serum albumin; Dextran sulfate; Heparin; Polyanion–protein complexes; Compressibility; Specific volume; Differential scanning calorimetry. * Corresponding author. Tel.: +421 2 60295683; fax: +421 2 65426774; e-mail: hianik@fmph.uniba.sk