ABC transporters Sav1866
The function of the ABC (ATP-binding-cassette) transporters is to release substances that are foreign to the organism from the inside to the outside of the cell. For example, MDR1 and MDP1 proteins of human contribute to the multidrug-resistance of cancer cells by discharging anticancer drugs. Moreover, the bacterial LmrA protein is involved in drug tolerance, and this is a major factor which causes a bacterial multidrug-resistant problem. Thus, it is very important to study these ABC transporter proteins. ABC transporters are membrane-bound proteins. That basic architecture consists of the transmembrane domain (TMD:Trans-membrane-domain) and the ATP binding domain (NBD:Nucleotide-binding-domain) located in the cytoplasm side. This protein exists as a dimer having two ATP binding site. A cavity is formed between the two TMDs to transports substances. The ATP binding region is well conserved. This means that most of the ABC family proteins have a common ATP hydrolysis and substance transport mechanism. However, despite extensive studies, a detailed understanding of the substrate transport process has remained elusive. In order to solve the mechanism, Dawson and others determined the structure of S. aureus Sav1866 which is an ABC transporter protein. Sav1866 shows significant sequence similarity to human ABC transporters that includes MDR1 and TAP1/TAP2. Based on the model suggested by Dawson, ABC transporter takes two conformations. First, before substance binding, the structure takes inward-facing conformation (open toward the inside of the cell). With substances binding to NBD, ATP also bind to NBD. Then, conformational change occurs, it becomes outward-facing conformation (open toward the outside of the cell), and the substance is discharged out of the cell. Next, if ATP is hydrolyzed, the structure change to inward-facing again that allows substance to access the protein. The structure determined by Dawson this time is an outward-facing conformation. In order to understand more detailed mechanism, we need to determine the structure of inward-facing conformation.
The structure determined this time is Sav1866 of Staphylococcus-aureus origin. (Fig.1). This protein forms a dimer coupled to two ATP. Each subunit is divided into TMD (1-320) which is a transmembrane domain, and NBD (337-578) which is an ATP binding domain on the side of cytoplasm. TMD consists of six transmenmrane alpha helices, and they are connected with three ECLs(loop outside a cell) and two ICLs (intracellular loop). Each ICL forms short αhelix and prominent about 25A to cytoplasm. A total of 12 alpha helices of TMDs are divided into two bunches, and are opened to the cell outside like two wings. A cavity formed between the wings is exposed to the extracellular space, but it is not connected with cytoplasm side. This feature shows that this structure is an outward-facing conformation. The two ATP binding sites exist in the interface of NBD-NBD binding, and ATP is firmly sandwiched by both of NBD. (Fig.2). Conformational change by binding of ATP or its hydrolysis in this region is transmitted to TMDs. TMD has interactions with the ATP binding site of NBD through the ICL1/ICL2. The two ICLs have a role of transmitting the conformational change to TMD.
Before the structure was solved, it was predicted that each TMD interacted with only NBD of own subunit. (Fig.3 left). However, in the observed structure, TMDs of Sav1866 take the twisted structure ; Each TMD interact with NBD of both subunits. (Fig.3 right). This cross-bonding pattern may be the feature of an outward-facing structure of ABC transporter.
Protein Data Bank (PDB)
Dawson, R.J.P. Locher, K.P.; "Structure of a bacterial multidrug ABC transporter."; Nature; (2006) 443:180-185 PubMed:16943773.
author: Jun-ichi Ito