Team:OUC-China/miniToe Family

Team OUC-China: Main

miniToe Family

1. Background

1.1 The Four Keys in miniToe System

The wetlab members give us four important sites, Gln104, Tyr176, Phe155, His29, which play import roles in binding and cleavage in protein Csy4. Considering 20 kinds of amino acids, we have 80 mutants to explore and choose if we only have one site mutated.

Before we begin to design the protein mutants, we first looking into the working process of miniToe structure to find that which are the most important keys in our system.

Fig.1-1 The working process of miniToe system

All the reaction happened in our first system, miniToe, can be described chronologically by following five main steps.
(1)The miniToe structure is produced and accumulated.
(2)The Csy4 is produced with IPTG induced.
(3)The Csy4 binds to the miniToe structure and form the rm the Csy4-miniToe complex
(4)The Csy4 cleavage the special site and divide the miniToe structure into two parts: the Csy4-crRNA complex and the mRNA of sfGFP.
(5)The sfGFP is produced.
From the description above, we can get four key problems in our system to make sure that our system can work successfully:
(1)Does the Csy4 dock correctly with the miniToe structure (hairpin)?
(2)How about the binding ability between the Csy4 and miniToe structure (hairpin)?
(3)How about the cleavage ability between the Csy4 and miniToe structure (hairpin)?
(4)Does crRNA release from the RBS?
The most impressive way to explore four problems is to model our system at the atom level by molecular dynamics. And there are lots of work in exploring the Csy4-RNA complex by molecular dynamics.

1.2 Molecular Dynamics

Molecular dynamics (MD) is a computer simulation method for studying the physical movements of atoms and molecules. The atoms and molecules are allowing to interact for a fixed period of time, giving a view of the dynamic evolution of the system. In the most common version, the trajectories of atoms and molecules are determined by numerically solving Newton's equations of motion for a system of interacting particles, whose forces between the particles and their potential energies are often calculated using interatomic potentials or molecular mechanics force fields.
To a system which consists of molecule or atoms, the total energy of a system includes kinetic energy and potential energy，which can be describe by the formula below: $E = {E_{kin}} + U$
where donates the kinetic energy and donates the potential energy.
In a molecule system, the total potential energy can be calculated by adding the 、 bond stretching potentials energy 、angle bending potentials energy 、torsion angle potentials energy 、out-of-plane potentials energy and some other cross effect together，which also can be describe by the formula below: $U = {U_{nb}} + {U_b} + {U_\theta } + {U_\phi } + {U_\chi } + {U_{cross}}$
This formula above also called the force field in molecular dynamics’ theory. There are many force field in the world that based on the statistical thermodynamics and empirical result. In the research of the protein and nucleic acid, the Amber force field is one of the best force field in the world. So we choose Amber as our force field and the formula of this field show below: \[\begin{array}{l} E = \sum\limits_{bond} {{K_b}{{({r_{ij}} - {r_0})}^2}} + \sum\limits_{angle} {{K_\theta }{{(\theta - {\theta _0})}^2}} + \sum\limits_{dihedral} {\frac{{{K_\phi }[1 + \cos (n\phi - {\phi _0})]}}{2}} \\ + \sum\limits_{impr} {\frac{{{K_\chi }[1 + \cos (n\chi - {\chi _0})]}}{2}} + \sum\limits_{nobond} {{\varepsilon _{ij}}\left[ {{{\left( {\frac{{R_{ij}^0}}{{{R_{ij}}}}} \right)}^{12}} - 2{{\left( {\frac{{R_{ij}^0}}{{{R_{ij}}}}} \right)}^6}} \right]} + \sum\limits_{nobond} {\frac{{{q_i}{q_j}}}{{{R_{ij}}}}} \end{array}\]
The items in the formula refers to bond stretching term、angle bending potentials、dihedral angle potentials、put of plane angle potentials、improper dihedral angle potentials、Van Der Waals interaction and Coulombic interaction terms in order.
Now considering the system which contains of consists of molecule or atoms in the classic mechanics, the atom or molecule can be characterized as follow:
The position of atom is , the speed of atom is , the acceleration of atom is , . After the integral operation, we can get two formula of and :