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Kinetics of D-amino...
Kinetics of D-amino acid incorporation in translation
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- Liljeruhm, Josefine (author)
- Uppsala universitet,Molekylärbiologi
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- Wang, Jinfan, 1989- (author)
- Uppsala universitet,Molekylärbiologi
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- Kwiatkowski, Marek (author)
- Uppsala universitet,Molekylärbiologi
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- Sabari, Samudra (author)
- Uppsala universitet,Molekylärbiologi
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- Forster, Anthony C. (author)
- Uppsala universitet,Molekylärbiologi,Anthony Forster
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(creator_code:org_t)
- 2019-01-16
- 2019
- English.
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In: ACS Chemical Biology. - : American Chemical Society (ACS). - 1554-8929 .- 1554-8937. ; 14:2, s. 204-213
- Related links:
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
Subject headings
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- Despite the stereospecificity of translation for l-amino acids (l-AAs) in vivo, synthetic biologists have enabled ribosomal incorporation of d-AAs in vitro toward encoding polypeptides with pharmacologically desirable properties. However, the steps in translation limiting d-AA incorporation need clarification. In this work, we compared d- and l-Phe incorporation in translation by quench-flow kinetics, measuring 250-fold slower incorporation into the dipeptide for the d isomer from a tRNAPhe-based adaptor (tRNAPheB). Incorporation was moderately hastened by tRNA body swaps and higher EF-Tu concentrations, indicating that binding by EF-Tu can be rate-limiting. However, from tRNAAlaB with a saturating concentration of EF-Tu, the slow d-Phe incorporation was unexpectedly very efficient in competition with incorporation of the l isomer, indicating fast binding to EF-Tu, fast binding of the resulting complex to the ribosome, and rate-limiting accommodation/peptide bond formation. Subsequent elongation with an l-AA was confirmed to be very slow and inefficient. This understanding helps rationalize incorporation efficiencies in vitro and stereospecific mechanisms in vivo and suggests approaches for improving incorporation.
Subject headings
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinsk bioteknologi -- Medicinsk bioteknologi (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Medical Biotechnology -- Medical Biotechnology (hsv//eng)
Publication and Content Type
- ref (subject category)
- art (subject category)
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