scholarly journals Loss of function from widely distributed, synonymous mutations at single codons

2019 ◽  
Author(s):  
Kritika Gupta ◽  
Jyothi Prabha ◽  
Soumyanetra Chandra ◽  
Shruti Khare ◽  
Sonali Vishwa Mohan ◽  
...  
Keyword(s):  
Rna Structure ◽  
Sequence Variation ◽  
Saturation Mutagenesis ◽  
Loss Of Function ◽  
Protein Ratio ◽  
Single Base ◽  
Natural Sequence ◽  
Synonymous Mutations ◽  
Natural Sequence Variation

AbstractMutational tolerance inferred from laboratory-based mutational studies is typically much higher than observed natural sequence variation. Using saturation mutagenesis, we show that the ccdA antitoxin component of the ccdAB toxin-antitoxin system is unusually sensitive to mutation with over 60% of mutations leading to loss of function. Multi-base synonymous mutations at a codon display enhanced propensity to show altered phenotypes, relative to single-base ones. Such mutations modulate RNA structure, leading to altered relative translation efficiencies of the two genes in the operon, and a CcdA:CcdB protein ratio below one. These insights were used to predict and experimentally validate synonymous mutations that lead to loss of function in the unrelated relBE operon as well as the lacZ gene. Thus, synonymous mutations can have significant phenotypic effects, in the absence of overexpression or extraneous reporters. More generally, proteins are likely more sensitive to mutation than inferred from previous saturation mutagenesis studies.

scholarly journals Codon optimality is the primary contributor to the exceptional mutational sensitivity of CcdA antitoxin in its operonic context

2022 ◽  
Author(s):  
Soumyanetra Chandra ◽  
Kritika Gupta ◽  
Shruti Khare ◽  
Pehu Kohli ◽  
Aparna Asok ◽  
...  
Keyword(s):  
Large Fraction ◽  
Point Mutations ◽  
Saturation Mutagenesis ◽  
Loss Of Function ◽  
Active Site Residues ◽  
Synonymous Mutations ◽  
Protein Levels ◽  
Intrinsically Disordered ◽  
Usage Frequency

Deep mutational scanning studies suggest that single synonymous mutations are typically silent and that most exposed, non active-site residues are tolerant to mutations. Here we show that the ccdA antitoxin component of the E.coli ccdAB toxin-antitoxin operonic system is unusually sensitive to mutations when studied in the operonic context. A large fraction (~80%) of single codon mutations, including many synonymous mutations in the ccdA gene show inactive phenotypes that are correlated with the E.coli codon usage frequency but retain native-like binding affinity towards cognate toxin, CcdB. Therefore, the observed phenotypic effects are largely not due to alterations in protein structure or stability, consistent with the fact that a large region of CcdA is intrinsically disordered. In select cases, proteomics studies reveal altered ratios of CcdA:CcdB protein levels in vivo, suggesting that these mutations likely alter relative translation efficiencies of the two genes in the operon. We extend these results by predicting and validating single synonymous mutations that lead to loss of function phenotypes in the relBE operon upon introduction of rarer codons. Thus, in their native context, genes are likely to be more sensitive to both synonymous and non-synonymous point mutations than inferred from previous saturation mutagenesis studies.

scholarly journals Analysis of the Effect of Natural Sequence Variation in Tat and in Cyclin T on the Formation and RNA Binding Properties of Tat-Cyclin T Complexes

1999 ◽  
Vol 73 (7) ◽  
pp. 5777-5786 ◽  
Author(s):  
Paul D. Bieniasz ◽  
Therese A. Grdina ◽  
Hal P. Bogerd ◽  
Bryan R. Cullen
Keyword(s):  
Sequence Variation ◽  
Rna Binding ◽  
Transcriptional Activator ◽  
Target Specificity ◽  
Tat Protein ◽  
Natural Sequence ◽  
Rna Targets ◽  
Immunodeficiency Virus ◽  
Natural Sequence Variation ◽  
Ltr Promoter

ABSTRACT The biological activity of the human immunodeficiency virus type 1 (HIV-1) Tat (Tat1) transcriptional activator requires the recruitment of a Tat1-CyclinT1 (CycT1) complex to the TAR RNA target encoded within the viral long terminal repeat (LTR). While other primate immunodeficiency viruses, such as HIV-2 and mandrill simian immunodeficiency virus (SIVmnd), also encode Tat proteins that activate transcription via RNA targets, these proteins differ significantly, both from each other and from Tat1, in terms of their ability to activate transcription directed by LTR promoter elements found in different HIV and SIV isolates. Here, we show that CycT1 also serves as an essential cofactor for HIV-2 Tat (Tat2) and SIVmnd Tat (Tat-M) function. Moreover, the CycT1 complex formed by each Tat protein displays a distinct RNA target specificity that accurately predicts the level of activation observed with a particular LTR. While Tat2 and Tat-M share the ability of Tat1 to bind to CycT1, they differ from Tat1 in that they are also able to bind to the related but distinct CycT2. However, the resultant Tat-CycT2 complexes fail to bind TAR and are therefore abortive. Surprisingly, mutation of a single residue in CycT2 (asparagine 260 to cysteine) rescues the ability of CycT2 to bind Tat1 and also activates not only TAR binding by all three Tat-CycT2 complexes but also Tat function. Therefore, the RNA target specificity of different Tat-CycT1 complexes is modulated by natural sequence variation in both the viral Tat transcriptional activator and in the host cell CycT molecule recruited by Tat. Further, the RNA target specificity of the resultant Tat-CycT1 complex accurately predicts the ability of that complex to activate transcription from a given LTR promoter element.

scholarly journals KinView: a visual comparative sequence analysis tool for integrated kinome research

2016 ◽  
Vol 12 (12) ◽  
pp. 3651-3665 ◽  
Author(s):  
Daniel Ian McSkimming ◽  
Shima Dastgheib ◽  
Timothy R. Baffi ◽  
Dominic P. Byrne ◽  
Samantha Ferries ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Sequence Variation ◽  
Comparative Sequence Analysis ◽  
Analysis Tool ◽  
Post Translational Modifications ◽  
Natural Sequence ◽  
Comparative Analyses ◽  
Comparative Sequence ◽  
Sequence Analysis Tool ◽  
Natural Sequence Variation

KinView enables both experts and novices to perform comparative analyses of cancer variants in the context of natural sequence variation and post-translational modifications across evolutionary groups of kinases.

Effects of natural sequence variation on symptom induction by citrus viroid III

1999 ◽  
Vol 134 (1) ◽  
pp. 73-80 ◽  
Author(s):  
R A OWENS ◽  
S M THOMPSON ◽  
P A FELDSTEIN ◽  
S M GARNSEY
Keyword(s):  
Sequence Variation ◽  
Natural Sequence ◽  
Natural Sequence Variation
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