Table 2.
A comparison of progeny phage output from a M13mp7L2 vector containing [+ta]-B[a]P-N2-dG as determined in E. coli carrying amino acid replacements at the roof-amino acid I38 in UmuC(V)1
| I38 | β | γ1 | γ2 | Yield | Protein Level3 |
|---|---|---|---|---|---|
| wt | -CH | -CH2CH3 | -CH3 | 100% (16) | 100% (3.9) |
| I38G | -H | -H | -H | 10% (2.9) | 48% (11) |
| I38A | -CH3 | -H | -H | 19% (4.0) | 99% (17) |
| I38C | -CHs | -SH | -H | 40% (16) | 130% (20) |
| I38V | -CH | -CH3 | -CH3 | 138% (9.9) | 95% (11) |
| I38M | -CH2 | -CH2SCH3 | -H | 12% (3.5) | 101% (20) |
| I38F | -CH2 | -Ph | -H | 20% (3.2) | 79% (22) |
| I38H | -CH2 | -Im | -H | 12% (4.5) | 140% (11) |
| I38L | -CH3 | -C3H72 | -H | 12% (3.0) | 119% (21) |
| I38S | -CH2 | -OH | -H | 16% (2.0) | 77% (13) |
| No UmuC | 2.9% (0.9) | <1% |
In parallel, a M13mp7L2 vector containing either [+ta]-B[a]P-N2-dG or no adduct was transformed into E. coli strains containing either wild type UmuC(V) (I38-wt) or various I38-mutants (e.g., I38G), or a cell with no UmuC (ΔumuC/D). The amino acid R-group is classified according to the atoms at the β-position, or the atoms attached to the β-carbon: γ1-only for straight-chain R-groups, or γ1- and γ2- if the β-carbon is branched. Progeny phage vector output (a measure of lesion bypass efficiency) was determined via plaque count (see text). Values (in percent) are the average of at least three experiments with standard deviation in parenthesis.
The amino acid leucine branches, but at the γ-position.
Protein level as determined by Western blotting. Values (in percent) are the average of at least three experiments with standard deviation in parenthesis.