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How a DNA polymerase Clamp Loader Opens a Sliding Clamp

            Deoxyribonucleic acid commonly called DNA is the basis if human life since it is the material that literally runs our body. DNA forms every cell in the body through several processes such as replication and translation. These processes result in the formation of new cells in the body or the repair of some old or damaged ones. DNA is made up of protein in the body and thus the processes involved require protein, as well. Replication is one of the most important processes that take place in the body, and it refers to the copying of the DNA strands. Replication provided the basis of the element of biological inheritance of traits and even diseases termed as hereditary. The process of replication depends on substances called multi-protein replicases.

They enable the copying of DNA to proceed at a faster rate and processivity. Replicases are examples of enzymes that DNA utilizes in order to accelerate the processes of replication in the body. Other such enzymes include the likes of ligases, helicases and polymerases. Every enzyme is formulated specifically to their function, and they, therefore, depict enzyme specificity. DNA occurs in a double strand and for it to undergo replication it needs to unwind. This is to enable the copying of both strands to take place effectively. After it is unwound, it forms a replication fork where the process takes place on the individual strands. The replicase involved in the replication process has several sub-units, but the most functional one in the case of replication is the polymerase sub-unit.

This sub-unit is attached to a ring-like sliding clamp that facilitates the polymerase to engage and disengage from the DNA repeatedly without undergoing dissociation from the replication fork. The use of sliding clamps is a common phenomenon, and both the eukaryotic and prokaryotic organisms use it. Eukaryotes are organisms whose cells are enclosed by a cell membrane while prokaryotes have no covering for their cells. However, the sliding clamps cannot load the DNA on impulse since they are closed circles and have no opening for the clamps. In order to load the DNA strand for replication to take place, they use ATP dependent complexes to create an opening for them to load on the DNA. ATP refers to adenosine triphosphate that acts as a source of energy for most processes taking part in the body.

These complexes called clamp loaders have a lot of energy stored, and they can part the closed ring and enable the replication process to begin. During the loading of the polymerase, it has to be loaded in the correct orientation in order to facilitate the required process to also take place correctly. Clamp loaders belong to the AAA+ super family that comprises of the adenosine triphosphates (ATpases). The main function of this family includes the ability to disassemble, helicases activity and motor activity. As opposed to the other members of the family, the clamp loaders have five sub units rather than the ordinary six contained in the rest of the family members. The lack of this one subunit is fundamental in their function as it enables them to be recognized by the primer templates.

ATP’s major role in the functioning of the clamp loaders is to stimulate the formation of AAA+ molecules in a spiral orientation. This enables them to recognize the DNA in the central region of the twirl they have formed. Some research had also signified that the ATP makes it possible for the loader to bind and untie the sliding clamp thus facilitating the replication to take place. DNA binding causes the hydrolysis of the ATP, which results in the liberation of the closed clamp present on the DNA. From research, it was concluded that DNA and the clamp loader have similar structures in both the complexes obtained from the first crystal. However, its contact with the lattice results in detachment of the loader though it is partial from the clamp if it was in a closed clamp.

The process of untying the clamp usually entails a switch from a ring-like structure that is planar and close to another open lock that is right handed. Their continued interactions result in maintaining the released state of the clamp. The clamp unlocks just as DNA does; the DNA is double-stranded and helical. Inside the complex, the closed clamp then positions the DNA appropriately for the required process to take place. The clamp that is now closed makes restricted contact with the DNA. Their interactions will include ion pairings between the side chains and the phosphates of DNA only. The open clamp takes up a spiral conformation that is identical to that of DNA. The sub-units of the clamp are associated with the other one by rotations about the axis in line with the axis of the DNA strand.

The clamp’s sideway opening results in the closest domains retracting from the DNA strand. It is probable that the clamp opening extent may cause the closing of loader complex before the DNA binding is completed. This is another reason why ATP is required as it enhances the stability of this complex, therefore, enables the binding process to be completed as required. The duplex region of the clamp loader is located in the inner chamber, which makes it oriented to the central pore. The template strand is the closest contact that the loader and the DNA have. The loader causes the conversion of the template strand from the B-form to the A-form. This conversion leads to an enlargement of the groove in order to support the incoming modules.

The template is then converted back into the B-form whereby the hydrolysis of ATP takes place. It is proposed that the hydrolysis of ATP results in the change, in the orientation of the clamp ladder while maintaining minimal interference to the contact points of DNA and those of the clamp. The ATP is hydrolyzed into its constituent components, which leave the complex as phosphate ions in two stages as three different ions. This results in the emergence of a clamp that is loaded with the template DNA strand. The clamp then detaches from the clamp loader which is then recycled in the body where the process takes place again. The clamp contains the template strand, which has already undergone replication in the cell.

The process by which a DNA Polymerase Clamp Loader unlocks a sliding clamp is often considered complicated. This is attributed to the various elements involved in the process and the numerous steps that culminate in the completion of this process. However, without the intricacies of the details is a simple procedure. It includes several small steps such as the binding of ATP in order to result in the required spiral conformation. A template then moves through the formed complex through the gaps. After the binding takes place in the interior chamber, the hydrolysis of the ATP is activated. The continuation of the hydrolysis leads to the detachment of the clamp from the clamp loader. After the completion of their process, the formed template may be used in repair or the formation of new cells. This process has to take place in the specified conditions or it may result in the formation of defective DNA that may result in congenital anomalies. It is, therefore, critical that this process takes place, as it is required.

Reference

Kelch, A. Brian et al. (2011). How a DNA polymerase Clamp Loader Opens a Sliding Clamp Science, Volume 10, pages 1-7.