Interaction of synthetic biomimetic nano-hydroxyapatite and nano-hydroxyapatite/alendronate with myoglobin

The interaction of proteins with solid surfaces is not only a fundamental phenomenon but is also key to several important and novel applications. In the biomaterials field, protein adsorption is the first step in the integration of an implanted device or material with tissue. In nanotechnology, protein–surface interactions are pivotal for the assembly of interfacial protein constructs, such as sensors, activators and other functional components at the biological/electronic junction. Fundamentally, the interaction of proteins with surfaces involves both protein binding and at least partial unfolding; these studies may therefore increase our knowledge of protein biophysics in general. Synthetic hydroxyapatite Ca10(PO4)6(OH)2 (HA) nanocrystals represent an elective material for bone substitutes. HA nano-crystals are similar to the mineral component of bone as composition and attempt to mimic the structural and morphological features of natural bone crystals by their nanosize, blade-like shape, low degree of crystallinity and high surface reactivity[1]. Bioactivity of a bone substitute material can be evaluated through its chemical reactivity with the physiological environment. In this sense the interaction between proteins and HA has received special attention because it can represent a model to investigate the way by which biomimetic materials communicate. Moreover, the use of probe proteins is very important to characterize the surface of substrate in wet conditions. For this purposes, myoglobin (from horse heart) (Mb), the oxygen binding protein in tissues, is a useful model to test as adsorbate. We investigated the isothermic kinetic adsorption of Mb onto different hydroxyapatites at several pH. In order to better understand this interaction, the HA/Mb has been explored by transmission electron microscopy (TEM), Raman spectroscopy and infrared spectroscopy (FT-IR). UV and CD spectroscopy investigations on the Mb solutions exchanged from the nanocrystals surface have given contributions towards understanding modifications in the protein structure due to the interaction with substrate.Chemical conjugation of bisphosphonates (BP’s) to HA is an effective means to impart it fine-tuned bioactivity[2]. This modification on the apatitic surface can be tested through its effect on the affinity towards proteins with respect to unloaded apatite. The first step is the binding of alendronate with HA and the second is the study of the adsorption of Mb onto this conjugate compared on its adsorption with unfunctionalized HA. The results show that Mb can not be completely adsorbed by the conjugates because the binding sites of the support are partially saturated by alendronate which also avoid the secondary structure modification of Mb. This study could improve our knowledge in the field of biomaterials science, in particular to fine tune a bone-specific drug delivery device and to test HA as a new support for enzyme immobilization.