Adenylate Kinase
Created by Julie Yee
Adenylate kinase (1NKS) from Sulfolobus acidocaldarius plays a key role in cellular energy metabolism. Adenylate kinase is involved in regulating phosphorolation, transferring phosphate groups from adenosine triphosphate (ATP) and other high-energy molecules to substrates (11). Adenylate kinase is a critical enzyme found in all cells to date, and is of special importance in cells that have a high ATP turnover rate (10, 1). It regulates the equilibrium between phosphorolation states and the recovery of adenosine monophosphate (AMP) by acting as a catalyst for ATP synthesis (6).
Although most members of the nucleoside monophosphate (NMP) family exist physiologically as a monomer, archaeal adenylate kinase exists physiologically as two asymmetric trimers (10). The subunits of the two trimers A-B-C(AMP) and D(AMP)-E-F(AMP, ADP) are each composed of 194 amino acid residues divided into three domains (11). The CORE domain consists of rigid β-sheets and contains the ATP binding site. The NMPbind domain consists of flexible α-helices involved in AMP binding. The LID domain consists of flexible α-helices involved in ATP and phosphoryl transfer (7). The molecular weight of adenylate kinase is 126,572.25 Da and the isoelectric point (pI) is 9.15, as provided by the Expasy Database (2). The secondary structure of adenylate kinase is 51% helical (8 helices) and 19% β-sheet (7 strands). It also has 6 β-turns and several bends (11).
The homology between adenylate kinase from Sulfolobus acidocaldarius, a thermophile, and adenylate kinases from different organisms is restricted to only 44 amino acid residues composing the central β-sheet and the phosphate binding loop (P-loop) surrounding the β-phosphate of ATP in the F subunit (7, 10). This P-loop is conserved in a variety of ATP-binding proteins across different organisms (7, 11). In addition, studies have shown that the removal of these N-terminal residues causes the protein to undergo a large conformational change into a more compact form (1). The sequence and structural conservation of this P-loop region across many NMP proteins demonstrate that the structure of adenylate kinase is related with its function.
The primary and tertiary structure of adenylate kinase is linked to its function. Adenylate kinase has two ligands, AMP and ADP. AMP is a monomer in RNA synthesis, and is composed of a phosphate group, ribose, and adenine. The AMP ligand binds to the enzyme in the NMPbind domain (11). In subunits C, D, and F, the adenine is held in place between Met-41 and Ile-96. The ribose is hydrogen bonded to the active site by Gln-69, Thr-92, and Gly-105. Thr-31 also forms a hydrogen bond with the ligand and rotates with it to induce closure (5). The active site residues Asn-36, Arg-54, His-93, and Arg-139 also change their conformation to accommodate and bind the AMP phosphate (11). ADP is used as an intermediate carrier of cellular energy, and is composed of a pyrophosphate group, ribose, and nucleoside adenine. The ADP molecule can easily bind to the CORE domain in the F subunit of adenylate kinase. The ADP adenine is held in place between Arg-132 and Pro-182, and is also hydrogen bonded to adenylate kinase’s Gly-180 and Asn-177. During phosphorolation, the magnesium ion is hydrogen bonded by Asp-91 (11). In addition, during catalysis, the NMPbind and LID domains close over the bound substrate and undergo a conformational change by moving closer to the central, compact, and rigid CORE (1, 7, 8).
Sequence and structural similarities between proteins can often indicate similar functions. PSI-BLAST is a query program that can use the amino acid sequence of a protein to search through an online database for other proteins of similar primary structures. A PSI-BLAST search was performed; however, the results with E values low enough to indicate significant similarity to adenylate kinase in Sulfolobus acidocaldarius were other forms of adenylate kinase expressed in other organisms. Proteins with an E value of less than 0.5 are sufficiently similar to the protein of interest. Adenylate kinase in Methanococcus Maripaludis (3H86) had an E-value of 4e-30, indicating that its primary sequence is significantly similar to that of the enzyme in Sulfolobus acidocaldarius (3). The Dali Server is another online query tool that can search for proteins with a similar tertiary structure to the protein of interest. Proteins with a Z score higher than 2 are sufficiently similar to the protein of interest. The Dali Server found that adenylate kinase in Methanococcus Maripaludis had a Z-score of 13.7, which indicates significantly high tertiary structure similarity to thermophilic adenylate kinase (4). Specific examples of tertiary structure similarity include the trimeric form and conservation of the central β-sheets, LID domain, and the P-loop at the ATP-binding site (9). Adenylate kinase regulates phosphorolation in both Sulfolobus acidocaldarius, a thermophile, as well as in Methanococcus Maripaludis, a methanogenic mesophile. To accommodate medium temperatures, mesophillic adenylate kinase has a rigid central trimer configuration. To accommodate higher temperatures, the thermostable adenylate kinase has an even stronger configuration enforced by the central β-sheet CORE (1, 9, 10). Although the two organisms differ in their optimal physiological temperature, adenylate kinases in both organisms have similar functions. This finding confirms the thesis that similar tertiary structures and conservation of the sequence at active-site regions may indicate similar functions.
Adenylate kinase has the important biological function of regulating cellular energy metabolism. There is conservation of residues at the active sites in many organisms, including Sulfolobus acidocaldarius and Methanococcus Maripaludis. The comparison of the sequence and tertiary structures of adenylate kinase in different organisms demonstrates that an analysis of the sequence and tertiary structure of the protein can indicate its function.