Figure 1.
Crystal structure of staphylococcus aureus IleRS-tRNA complex (Protein Data Bank) [16].
tRNA is shown in orange, CP1 domain in green, Rossmann fold in red, and rest of the protein is represented in blue.
Figure 2.
Schematic representation of one of the two kinetic proofreading mechanisms (mechanism I) proposed by Fersht.
Here formation of Val-tRNAIle is the rate limiting step (k = 1.2 s−1) and it is followed by rapid hydrolysis of the complex.
Figure 3.
An alternate mechanism of kinetic proofreading (mechanism II) proposed by Fersht.
In this mechanism change of enzyme conformation is the slowest step. The overall kinetic involves hydrolysis in two steps.
Figure 4.
The proposed kinetic scheme for isoleucine.
The symbols are described in the text.
Figure 5.
The proposed kinetic scheme for valine.
The symbols are described in the text.
Figure 6.
The first passage time distribution for the aminoacylation of tRNAIle by isoleucine in single molecular reaction, under single molecular catalytic conditions.
Figure 7.
The computed first passage time distribution for the aminoacylation of tRNAIle by valine in single molecular reaction.
The inset shows the time dependence at short time where the distribution shows an initial sharp rise followed by a slow decay.
Figure 8.
Time dependence of AMP concentration generated by ATP hydrolysis for different tRNAIle concentrations during synthesis of Val-tRNAIle.
Figure 9.
tRNAIle concentration dependence of steady state rate of ATP hydrolysis for (A) valine and (B) isoleucine.
The insets show their corresponding behaviors in inverse scales. Note the deviation from Michaelis-Menten behavior for isoleucine.
Figure 10.
tRNAIle concentration dependent steady state rate of ATP hydrolysis for Val in pre- and post transfer processes.
The fraction of pre-transfer hydrolysis with respect to the overall ATP hydrolysis is also shown with a blue dashed line. The relative contribution of pre-transfer editing converges to a finite value (∼9%) at high tRNAIle concentration.
Figure 11.
Time dependence of fraction of free enzyme and enzyme-tRNAIle complex at two different tRNAIle concentrations ([tRNAIle] = 1 nM and 100 nM) under multi-turnover condition in presence of valine.
The dashed and solid lines are for 1 nM and 100 nM concentrations of tRNAIle, respectively. Black and green lines indicate fraction of free enzyme whereas the red and blue lines indicate fraction of enzyme-tRNA complex.
Figure 12.
Amino acid concentration dependence of steady state rate of enzyme catalysis for isoleucine (black solid line) and valine (blue dashed line).
The rate of ATP hydrolysis for Val is shown by red dot-dashed line.
Figure 13.
Left panel: The time dependence of products (Ile-tRNAIle and Val-tRNAIle) when both isoleucine and valine are present in the system with equal amount.
Inset shows the linear increase of Val-tRNAIle during steady state. Right panel: Amino acid concentration dependence of overall editing (i.e., the ratio of steady state rates of Ile-tRNAIle and Val-tRNAIle formation in presence of equal amounts of Ile and Val).