(Back to
the menu - click here.)
“Accuracy of genetic
code translation by transfer RNAs
on the messenger RNA
programmed ribosome “
|
Date: |
Download-files: |
Time: |
|
Thursday, 25. Feb. 2016 |
Audio-only-Recording as MP3-File (smallest
possible size):
- Audio.mp3 (ca.30 Mb) ============================================ Video-Recording for any system with MP4-support:
- Video.mp4 (ca.270 Mb) |
15:15 – 15:20 |
Speaker :
Måns Ehrenberg (
Abstract :
Misreading
of messenger RNAs (mRNAs) by transfer RNAs (tRNAs)
on bacterial ribosomes has been intensely discussed
over the last fifty years, but quantitative information of the missense error spectrum remains scarce. It is known that aminoacyl-tRNAs (aa-tRNAs) enter
the ribosome in “ternary complex” (T3) with a protein factor, the GTPase EF-Tu. Depending on if the
anticodon base triplet of the tRNA
matches the codon base triplet a ternary complex is
accepted for GTP hydrolysis on EF-Tu or discarded in
the initial selection step of mRNA reading. After initial selection non cognate
tRNAs which have
"survived" the initial selection step is discarded with high
probability in a proofreading step, occurring after GTP hydrolysis (Thompson
and Stone, 1977; Ruusala et al., 1982). It is the
chemical potential difference between GTP and its hydrolytic products that
makes proofreading possible and determines its universal accuracy limits
(Ehrenberg and Blomberg, 1980). There has been
scientific consensus that genetic code translation errors predominantly come
from mRNA reading on the ribosome (0.0005 per codon)
and to a lesser extent from transcription of DNA to mRNA (0.0001 per base) and aminoacylation of tRNA to aa-tRNA (0.00001 per reaction).
We have
shown by biochemical experiments involving seven tRNAs
reading their cognate and all near-cognate codons
that initial selection of tRNAs spans over three
orders of magnitude from an intrinsic accuracy of about 100 to 100 000 (Zhang
et al., 2015). The initial selection accuracy is generally very high (>10
000), but there are a small number of initial error hot-spots involving middle codon position misreading by tRNAGlu
and third codon position misreading by tRNAHis. Very recently, we determined the total accuracy
(A=IF) for three of these tRNAs
and how it is partitioned between proofreading (F) and initial (I) selection
(Submitted manuscript). An amazing finding is that in the high accuracy range log(F) is a linear function of log(I) with a slope close to
two. In the low accuracy range, in contrast, log(F) is
constant as log(I) decreases further. Since the same discrimination parameters
are at work in both proofreading and initial selection it may seem paradoxical
that proofreading can remain at a high level and thus neutralize error hot
spots while initial selection plummets. To explain this, we propose that the
expressed fraction of the intrinsic accuracy in proofreading selection
increases and compensates for the reduction of intrinsic accuracy itself, as
revealed by decreasing initial selection. We point out that the slope close to
two in the log(F) versus log(I) plot most naturally
arises from two consecutive proofreading steps (Ehrenberg and Blomberg, 1980) rather than one, as one has hitherto
postulated.
We have,
finally, directly proved the existence of two consecutive proofreading steps by
engineering tRNAs with
different affinities to EF-Tu and studying how the
proofreading selection varies with varying aa-tRNA-EF-Tu
affinity (Submitted manuscript).
Thompson
RC, Stone PJ (1977) PNAS 1: 198-202
Ruusala
T, Ehrenberg M,
Ehrenberg
M, Blomberg C (1980) Biophys.
J 31: 333-358
Zhang J, Ieong KW, Johansson M, Ehrenberg M (2015) PNAS 112
9602-9607