| E.coli |
|
|
|
| Discriminator base (A73) |
Replaced with C, U or G |
Base preference: A>C>U>G |
(2) |
| |
tRNAser G73 replaced with A |
Conferred leucylation |
(5) |
| |
Replaced with C (ΔACVL1) |
Leucylation abolished |
This work |
| Acceptor stem |
G1 changed to A1 |
No effect |
(2) |
| |
C2:G71 changed to G2:C71 |
No effect |
|
| |
Leucine minihelix |
No aminoacylation |
This work |
| D-stem–loop |
U16 changed to A16 |
No effect |
(2) |
| |
A14 changed to C, U or G |
Decreased 100-fold |
|
| |
tRNAser G18G19 location altered |
Conferred leucylation |
|
| |
tRNAtyr G18G19 location altered |
Conferred leucylation |
|
| |
A15:U48 changed to G15:C48 |
Small reduction |
|
| |
Randomized for in vitro selection |
A15, G18, G19, A20a conserved |
(6) |
| |
|
G18:G19 fixed position |
|
| |
tRNAser G15:C48 changed to A15:U48 |
In vivo analysis: 82% suppression |
(5) |
| |
tRNAser G18G19 location altered |
100% leu insertion |
|
| |
D-stem–loop domain deleted (ΔACDL1–4) |
No aminoacylation |
This work |
| |
G18 changed to A18 (ΔACVL1) |
Aminoacylation abolished |
|
| |
G19 changed to A19 (ΔACVL1) |
Aminoacylation abolished |
|
| |
G18G19 changed to A18A19 (ΔACVL1) |
Aminoacylation abolished |
|
| Anticodon stem–loop |
G37 changed to U37 |
No effect |
(2) |
| |
U32 changed to C32 |
No effect |
|
| |
Anticodon domain deleted (ΔAC) |
Reduced 17-fold |
This work |
| Variable arm |
G45:C47I changed to C45:G47I |
No effect |
(2) |
| |
G47J changed to A47J |
Reduced 2-fold |
|
| |
Two base pair deletions |
Reduced 2-fold |
|
| |
Randomized for in vitro selection |
U48 conserved |
(6) |
| |
|
Variants are good substrates |
|
| |
47 nt inserted in tRNAser
|
Conferred leucylation |
(5) |
| |
Variable domain deleted (ΔVL) |
Reduced 125-fold |
This work |
| |
Anticodon stem–loop and variable domains deleted (ΔACVL1–3) |
Reduced significantly |
|
| T-stem–loop |
N59 randomized for in vitro selection |
No effect |
(6) |
| |
U55 changed to C55 (ΔACVL1) |
Aminoacylation abolished |
This work |
| |
C56 changed to U56 (ΔACVL1) |
Aminoacylation abolished |
|
| |
U55C56 changed to C55U56 (ΔACVL1) |
Aminoacylation abolished |
|
| Human (cyt) |
|
|
|
| Discriminator base (A73) |
U73 |
Low efficiency |
(4) |
| |
C73 and G73 |
Reduced aminoacylation |
|
| Acceptor stem |
C3:G70 inserted into tRNAser
|
Conferred leucylation 50-fold |
|
| |
A4:U69 inserted into tRNAser
|
|
|
| |
G5:C68 inserted into tRNAser
|
|
|
| |
Leucine minihelix |
No aminoacylation |
(7) |
| D-stem–loop |
Removed C20a |
Decreased 30-fold |
(24) |
| Variable arm |
Variable loop inserted into tRNAser
|
Conferred leucylation |
(24) |
| |
Variable loop replaced with 5 nt arm |
Abolished leucylation |
|
| T-stem-loop |
U50:A64 inserted into tRNAser
|
No effect |
(24) |
| |
G51:C63 inserted into tRNAser
|
No effect |
(24) |
| Yeast (cyt) |
|
|
|
| Discriminator base (A73) |
A73 (plus A35 and G37 in anticodon) inserted into tRNAser |
Conferred leucylation |
(3) |
| Anticodon stem–loop |
A35 and G37 |
Necessary identity determinants |
(3) |
