Conformational transitions of the three recombinant domains of human serumalbumin depending on pH

Human serum albumin (HSA) is a protein of 66.5 kDa that is composed of thre e homologous domains, each of which displays specific structural and functi onal characteristics. HSA is known to undergo different pH-dependent struct ural transitions, the N-F and F-E transitions in the acid pH region and the N-B transition at slightly alkaline pH, In order to elucidate the structur al behavior of the recombinant HSA domains as stand-alone proteins and to i nvestigate the molecular and structural origins of the pH-induced conformat ional changes of the intact molecule, we have employed fluorescence and cir cular dichroic methods. Here we provide evidence that the loosening of the HSA structure in the N-F transition takes place primarily in HSA-DOM III an d that HSA-DOM I undergoes a structural rearrangement with only minor chang es in secondary structure, whereas HSA-DOM II transforms to a molten globul e-like state as the pH is reduced. In the pH region of the N-B transition o f HSA, HSA-DOM I and HSA-DOM II experience a tertiary structural isomerizat ion, whereas with HSA-DOM III no alterations in tertiary structure are obse rved, as judged from near-UV CD and fluorescence measurements.