Serum Albumin Complexed with Lidocaine
Created by Rebecca Grande
Human serum albumin is a transport protein in the albuminoid superfamily, essential for binding and transporting several electronegative and hydrophobic organic compounds present in blood plasma and extravascular spaces, such as hormones, fatty acids, and drugs, which in this case include lidocaine (PDB ID 3JQZ). The albuminoid superfamily, comprised of serum albumin, alpha-feto protein, alpha-albumin, afamin, and vitamin-D binding (Gc) protein, contains three characteristic domains shared by these proteins (3). The conservation of these domains in all proteins of the superfamily indicates their similar function of serving as transport proteins for several different ligands. These domains consist of five or six internal disulfide bonds, and the family itself has highly conserved intron/exon organization (BLAST results).
Human serum albumin (HSA) is the most abundant protein in the cardiovascular system, comprising about 60% of plasma protein, and may also be found in extravascular spaces at low concentration (4). It directly controls the distribution of drugs throughout the body, as most drugs reach target tissues bound to HSA while they travel in plasma (6). HSA consists of a chain 586 amino acids in length. These residues are structured into 67% α helices (2). Being a part of the albuminoid superfamily, HSA contains three characteristic homologous domains, each of which has two subdomains (Protein Data Bank ID 3JQZ results). Each domain has a distinct helical folding pattern connected by flexible loops and they all have different capacities for binding fatty acids, hormones, and drugs, despite their internal structural symmetry (1). Domain I includes residues 5-197 (subdomain IA: 5-107, subdomain IB: 108-197). Domain II is made of residues 198-382 (subdomain IIA: 198-296, subdomain IIB: 297-382), and domain III consists of residues 383-569 (subdomain IIIA: 383-494, subdomain IIIB: 495-569).
Transport proteins all generally bind to a specific ligand necessary for a certain pathway, and they transfer this ligand to the active site in which it is needed. In the particular case for lidocaine, a local anesthetic drug, it must block sodium (Na+) channels in neurons responsible for signal propagation to relieve the sensation of pain. HSA is monomeric protein, but upon interaction with lidocaine, it becomes an asymmetric dimer, consisting of one serum albumin molecule and another serum albumin molecule bound to a single lidocaine unit (2). Domain I of the HSA molecule without lidocaine is oriented closer to the middle of the dimer than the HSA binding lidocaine. Lidocaine, a tertiary amine compound, binds to a low-affinity site, which still distributes the drug effectively due to the high concentration of HSA in the bloodstream. HSA has a certain number of high-affinity binding sites; for example, there are two specialized drug binding pockets within subdomains IIA and IIIA that anchor the substrate within the site using hydrogen bonds (5). Hydrophobic interactions, along with polar interactions at the surface of the protein, contribute to the high-affinity cavities found in subdomain IB, subdomain IIA, subdomain IIIA, and subdomain IIIB, which bind to aliphatic chains of cationic aliphatic detergents as well as drugs with anionic or electronegative features (2). These hydrophobic interactions are absent at the lidocaine-binding site. This low-affinity site is found on subdomain IB, in an area formed by residues subdomain I and III, both facing the central, interdomain crevice (2). Although the area has a lower affinity for binding, the crevice makes it easier to hold on to the lidocaine molecule. Lidocaine is able to interact electrostatically with the positively charged, tertiary amine nitrogen and carboxylate group of Asp-187 (2). The carbonyl group of lidocaine has a polar interaction with Lys-190, while the aromatic portion is involved in cation-π interactions in Arg-114 (2). This aromatic portion attracts Arg-114, displacing the amino acid, while Arg-186 is also displaced. The HSA-lidocaine complex causes domain III of HSA with no ligand to be more outplaced than the HSA unit bound to lidocaine, and domain I of HSA with no ligand becomes tilted towards the middle. Lidocaine also induces conformational changes in the structural features of Trp-214, which leads to a decreased polarity around it (2). These structural changes differ from those induced by fatty acids in high-affinity hydrophobic binding sites. Fatty acid binding in I-II and II-III interfaces results in rigid-body rotations relative to domain II, in which domains I and III pivot around the mid-point of the long inter-domain helices (1). Domain III is also further away from the central crevice than in the HSA-lidocaine complex and domain I is closer to the central crevice (2). Domains I and III do not rotate when HSA binds to lidocaine. The interactions of lidocaine with amino acids in subdomain IB stabilize the HSA-lidocaine structure. The positively-charged residues in the binding site allow for this weak drug binding. Furthermore, there are no hydrophobic regions in this pocket to increase affinity for another particular molecule.
Once lidocaine is complexed with HSA, the protein carries the drug to the α-subunit of neuron sodium channels responsible for transmitting the sensation of pain. The aromatic portion of lidocaine must interact with phenylalanine in Na+ channels in order to bind with them. This is also facilitated by the electrostatic attraction of the tertiary amine nitrogen to glutamic acid. Because interactions with phenylalaline and glutamic acid of Na+ channels are essential for drug binding, lidocaine binds with arginine instead of phenylalanine and aspartate instead of glutamate. Once the protein comes close to Na+ channels, lidocaine transitions from a low-affinity site on HSA to that of a higher affinity at the Na+ channel. If there were any mutations that replaced the amino acids in subdomain IB (specifically arginine, aspartic acid, and lysine) with hydrophobic residues, lidocaine would not be able to bind, given its already weak nature to bind with HSA. The low-affinity of the binding site, and thus less specific selection for binding in this particular site, allows for investigation concerning what other drugs can be manufactures to be carried by human serum albumin. This knowledge may also be combined with information obtained from the binding of fatty acids to HSA, which have shown to influence the binding of other drugs to HSA (7). Conformational changes induced by fatty acid binding may assist in binding other ligands to HSA. Knowledge of both high-affinity and low-affinity binding will facilitate the design of drugs depending on whether they are meant to bind with HSA or to avoid binding to it altogether.