One of human serum albumin's most outstanding qualities is its ability to bind reversibly an incredible variety of ligands. HSA is the principal carrier of fatty acids that are otherwise insoluble in circulating plasma. It performs many other functions as well, such as sequestering oxygen free radicals and inactivating various toxic lipophilic metabolites such as bilirubin. The ligands of HSA can be separated into three categories: long chain fatty acids, metals, and small organic compounds.
Due to the incredible diversity of ligands bound by albumin, early researchers
saw ligand binding to HSA as nonspecific in nature and did not recognize
that there were discrete sites. Instead they envisaged the ligands
as randomly attached to the surface and envisioned HSA as some what like
a sponge. This uninteresting view of albumin has changed over the
past years, and now it is generally recognized that there are a small number
of distinct binding locations.
• LONG CHAIN FATTY ACID BINDING
HSA is recognized as the principle transport protein for fatty acids and other lipids that would otherwise be insoluble in the circulating plasma. Studies have implied that there are two high-affinity and four low-affinity long-chain fatty acid binding sites. There appears to be a general agreement that the major sites reside in domains I and III. Binding of fatty acid induces a dramatic conformational change which has both universal and local aspects. There is no clear consensus within the current scientific literature on the exact location of bound long chain fatty acids, except that their locations do not correspond to the sites of small organic compounds. Various studies have show that Arg-117, His-146, Lys-351, Lys- 475 and Lys-525 are involved in fatty acid binding.
MYRISTATE (LONG CHAIN FATTY ACID) - Click the button to see a representation of the albumin-myristate complex. The five myristates are colored white and the subdomains are labeled with different colors.
The relative shift associated with binding myristic acid can be seen by comparing the bound HSA to the left with the unbound below.
Notice how the blue area shifts. In the above figure, the vertical helix curves to the right and the horizontal helix is completely horizontal. When myristic is bound, shown to the right, the vertical helix bends to the right and the horizontal helix shifts upward. As a result, there is a larger angle between the two helices in the bound form.
• MYRISTATE BINDING AT A CLOSER LEVEL
While the universal changes in domain orientation are quite dramatic, the local effects of fatty acid binding on the protein are more subtle. These changes can be seen at a closer level. The purple region of the above picture is a myristic binding site. Below is a representation of this area unliganded. The amino acids Ser-287, Arg-257, Tyr-150, Leu-251 are shown.
Compare the unbound structure, below, with that of the bound structure to the left.
The binding of myristate at the interface between subdomains IA and IIA appears to change the rotation of domain I relative to domain II. The arrows indicate the direction of movement. Comparison of the two views shows how Leu-251 fills the hydrophobic portion of the pocket in the absence of bound fatty acid (above). When bound, the fatty acid displaces the area where the Leu-251 would be in the unbound form. Of the three side chains that are observed to hydrogen bond to the carboxylate group of myristate, only Tyr-150 is observed to significantly change conformation; the positions of Arg-257 and Ser-287 on IIA are altered only slightly. Binding of long chain fatty acids in general, is similar to that seen in myristate binding. Large conformational changes are seen universally and small subtle changes are seen locally.
• METAL BINDING
HSA has a high affinity for Cu (II), Ni (II), Hg (II), Cd (II), Au (I), and Ag (II), and weaker affinities for calcium and zinc. It possesses two major binding sites for metals and a new binding site has come to light recently.
CYS-34 BINDING SITE - Albumin is responsible for the largest fraction of free sulfhydryl (Cys-34) in blood serum and studies have shown that it is also the most reactive sulfhydryl. Cys-34 initiates dimer formation between HSA's and binds Au, Ag, Hg, Cd, and, to a lesser extent, Cu. Cys-34 is located in a crevice on the surface of the protein and the reactive sulfur is somewhat protected by several residues.
Click this button for the representation in spacefill. It appears as though it is inaccessable from the front of HSA.
This view shows that it is accessable from the top.
N TERMINUS BINDING SITE - Albumin possesses one distinct high-affinity site for Cu (II) and Ni (II) , which has been well characterized by several research groups. The binding of Cu by HSA was identified with the N terminus of HSA. They believed that the copper atom was coordinated by the N-terminal nitrogen, the next two peptide nitrogens, and the side group N of His-3. It was observed that the tripeptide glycylglycyl-L-histidine-N-methylamide, a functional analog of the N-terminal tripeptide of HSA, demonstrated similar copper binding chemistry.
Figure. Structure of the copper-binding peptide, glycylglycyl-L-histidine-N-methylamide
illustrating the coordination state and peptide conformation.
PROPOSED BINDING SITE FOR Cd (II) AND Zn (II) - Recent research has suggested HSA possesses a strong Cd (II) / Zn (II) binding site which does not involve the free thiol at Cys-34 or N-terminus. Data suggested that the thiolate sulfur of Cys-34 was not a ligand in either of the two strong Cd (II) / Zn (II) binding sites, since the affinity of Cd (II) and Zn (II) for bovine serum albumin, a close homolog of HSA, is unaffected by the blocking of Cys-34. Other work has has suggested the involvement of His residues in the binding of Cd (II) and Zn (II) to HSA. There seemed to be only one area within HSA in which two, or possibly three, His residues lie close in space: His-105 and His-247 in domain I, and His-247 from domain II.
Figure. Proposal for the shared Cd (II) and Zn (II) binding site in
HSA; containing His-105, His-146, His-247, and one or more carboxylate
ligands (Asp-249, Asp-107, or Asp-108). The "M" represents
either Cd (II) or Zn (II).
Click here for a respresentation of this. His-105, His-146, and His-247 are represented in color.