Circadian Clock Protein KaiC

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Contents 1 Introduction 2 KaiC - KaiA - KaiB System 3 KaiC Homohexameric Complex 4 KaiC Autophosphorylation Sites 5 KaiA <-> KaiC Interaction Site 6 KaiB <-> KaiC Interaction Site 7 Biological Importance

Introduction

Biological Circadian Clocks are self-sustaining oscillators that function on a rhythmic cycle of or around 24 hours. The are found in almost all organisms, the simplest of which are cyanobacteria, which have been extensively studied in order to determine the mechanism of the fine-tuned biological process of circadian rhythmicity. In most eukaryotes, a region of the brain called the superchiasmic nuclei detects light and dark cycles that relay the message to biological clock systems that maintain rhythmicity within the body. Conversely, cyanobacteria have a fairly small system comprised of three proteins: KaiC, KaiA, and KaiB. The system is based around the central protein KaiC which exhibits ATP binding, inter-subunit organization, a scaffold region for Kai protein complex formation, a location where critical mutations are found, and an evolutionary link to other well-known proteins (1).

KaiC - KaiA - KaiB System

KaiC is the central clock protein, which has autokinase and autophosphorylase activity. Yet in the presence of ATP, KaiC cannot perform it's autophosphorylation function. It requires two other proteins, KaiA and KaiB, the genes of which are found in the same cluster on the chromosome (1). Although KaiC phosphorylates itself, the presence of KaiA and KaiB are essential to rhythmicity. KaiA stimulates KaiC autophosphorylation, while KaiB antagonizes the process possibly by enhancing KaiC dephosphorylation. Even in the presence of high ATP, KaiB still prompts KaiC to dephosphorylate.

KaiC Homohexameric Complex

spinqualitylabelsall models Structure of KaiC (PDB entry 1TF7) Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image The structure of KaiC resembles a double donut formation with a central pore through the center. Conceptually, it is comprised of two subunits, which appear as the two donuts. Yet, technically, the protein divided perpendicular to the conceptual lateral subdivision, which can be visualized as six that form a barrel shaped structure. The entire molecule is made up of 519 amino acid residues, and spans 100 Angstroms in diameter. The waist of the molecule (the region between CI and CII) has a diameter of 62 Angstroms. The that runs through the molecule is wider at the CI region, 22 Angstroms, and narrows to a nearly that appears to be secured by six arginine residues. The basic, polar nature and physical divergence of this CII region in comparison to the CI end indicates the possibility of conformational change, such as closing and opening. (1) There are 12 molecules of bound at the interface between CI and CII. Three potential phosphorylation sites have been identified within 10 Angstroms of the ATP binding region.

KaiC Autophosphorylation Sites

The phosphorylation sites on the KaiC protein are essential to the system. This is because phosphorylation status corresponds to clock speed. The protein predominantly phosphorylates on threonine and serine residues, whose specific identification is not completely resolved. Nonetheless, three potential phosphorylation sites have been identified within 10 Angstroms of the ATP binding region in the CII domain (2). The key autophosphorylation site is T432. The process is believed to demonstrate a transfer of the gamma phosphate of ATP from one CII subunit to the T432 site on an adjacent subunit. A region of each hexamer that is notable regarding the phosphorylation of KaiC is the P Loop. This zone is recognized as site for binding and hydrolysis of ATP. Along with the T432 site, evidence shows a shuttling of phosphates between residues S431 and T426 of the P Loop. (difference between ATP binding and phosphorylation residues) (Key residues = T432, S431, T426) (show 3D image of these sites and highlight bonds/interactions)

KaiA <-> KaiC Interaction Site

(show 3D image of KaiC site for KaiA binding and highlight key residues in interaction -weak? strong? any ions in site? how does it stabilize autophosporylation?) KaiA binds to the interface of the two donut-shaped KaiC subunits, CI and CII. This area, known as the "waist" of the molecule

KaiB <-> KaiC Interaction Site

(show 3D image of KaiC site for KaiB binding and highlight key residues in interaction -weak? strong? ions in site? how does it stabilize dephosphorylation/destabilize phosphorylation/destabilize KaiA?)

Biological Importance

- Nearly all promoters in a cyanobacteria are under circadian control. [function is important to whole life cycle] (1)