Abstract:Compared to folded proteins, the sequences of intrinsically disordered proteins (IDPs) are enriched in polar and charged amino acids. Glutamate is one of the most enriched amino acids in IDPs, while the chemically similar amino acid aspartate is less enriched. So far, the underlying functional differences between glutamates and aspartates in IDPs remain poorly understood. In this study, we examine the differential effects of aspartate and glutamates in IDPs by comparing the function and conformational ensemble… Show more
“…The converse was also true, as replacing the relatively few D residues in the linker region with E residues (LR_DtoE) also reduced the helical content. However, LR_DtoE maintained more helicity than LR_E-toD, which is consistent with a recent study suggesting that glutamate helps maintain helicity, while aspartate helps maintain extended structures in IDPs (Roesgaard et al, 2022). This trend partially carried over to positively charged residues as well, with our LR_RtoK variant showing reduced helical content, while the helical content of the LR_KtoR variant remained more or less consistent with that of the wild-type linker region.…”
Section: Sequence Identity Of Charged Residues In the Cahs D Linker P...supporting
In order to survive extreme drying (anhydrobiosis), many organisms, spanning every kingdom of life, accumulate intrinsically disordered proteins (IDPs). For decades, the ability of anhydrobiosis‐related IDPs to form transient amphipathic helices has been suggested to be important for promoting desiccation tolerance. However, evidence empirically supporting the necessity and/or sufficiency of helicity in mediating anhydrobiosis is lacking. Here we demonstrate that the linker region of CAHS D, a desiccation‐related IDP from the tardigrade Hypsibius exemplaris, that contains significant helical structure, is the protective portion of this protein. Perturbing the sequence composition and grammar of the linker region of CAHS D, through the insertion of helix‐breaking prolines, modulating the identity of charged residues, or replacement of hydrophobic amino acids with serine or glycine residues results in variants with different degrees of helical structure. Importantly, correlation of protective capacity and helical content in variants generated through different helix perturbing modalities do not show as strong a trend, suggesting that while helicity is important it is not the only property that makes a protein protective during desiccation. These results provide direct evidence for the decades old theory that helicity of desiccation‐related IDPs is linked to their anhydrobiotic capacity.This article is protected by copyright. All rights reserved.
“…The converse was also true, as replacing the relatively few D residues in the linker region with E residues (LR_DtoE) also reduced the helical content. However, LR_DtoE maintained more helicity than LR_E-toD, which is consistent with a recent study suggesting that glutamate helps maintain helicity, while aspartate helps maintain extended structures in IDPs (Roesgaard et al, 2022). This trend partially carried over to positively charged residues as well, with our LR_RtoK variant showing reduced helical content, while the helical content of the LR_KtoR variant remained more or less consistent with that of the wild-type linker region.…”
Section: Sequence Identity Of Charged Residues In the Cahs D Linker P...supporting
In order to survive extreme drying (anhydrobiosis), many organisms, spanning every kingdom of life, accumulate intrinsically disordered proteins (IDPs). For decades, the ability of anhydrobiosis‐related IDPs to form transient amphipathic helices has been suggested to be important for promoting desiccation tolerance. However, evidence empirically supporting the necessity and/or sufficiency of helicity in mediating anhydrobiosis is lacking. Here we demonstrate that the linker region of CAHS D, a desiccation‐related IDP from the tardigrade Hypsibius exemplaris, that contains significant helical structure, is the protective portion of this protein. Perturbing the sequence composition and grammar of the linker region of CAHS D, through the insertion of helix‐breaking prolines, modulating the identity of charged residues, or replacement of hydrophobic amino acids with serine or glycine residues results in variants with different degrees of helical structure. Importantly, correlation of protective capacity and helical content in variants generated through different helix perturbing modalities do not show as strong a trend, suggesting that while helicity is important it is not the only property that makes a protein protective during desiccation. These results provide direct evidence for the decades old theory that helicity of desiccation‐related IDPs is linked to their anhydrobiotic capacity.This article is protected by copyright. All rights reserved.
“…Studies have shown that small differences in amino acid chemistries are of immense significance . As a result, Ser and Thr are not always interoperable, nor are Arg and Lys, ,, Glu and Asp, ,, Gln and Asn, or Phe and Tyr . Any interoperability is highly sequence- or context-specific .…”
Multivalent proteins and nucleic acids, collectively referred to as multivalent associative biomacromolecules, provide the driving forces for the formation and compositional regulation of biomolecular condensates. Here, we review the key concepts of phase transitions of aqueous solutions of associative biomacromolecules, specifically proteins that include folded domains and intrinsically disordered regions. The phase transitions of these systems come under the rubric of coupled associative and segregative transitions. The concepts underlying these processes are presented, and their relevance to biomolecular condensates is discussed.
“…Thus, of the proteins with lysine deserts longer than 200 residues (after Shannon filtering for low-complexity regions) in the yeasts S. cerevisiae and S. pombe , we found by manual curation that 7 out of 14 (50%) and 17 out of 35 (48%), respectively, are UPS components (Supplemental file 3). The previously described budding yeast lysine desert proteins Slx5 and San1 [ 17 , 18 ] were the highest scoring hits in S. cerevisiae (Supplemental file 3) [ 28 , 29 ]. Fig.…”
Section: Resultsmentioning
confidence: 99%
“…Accordingly, the lysine deserts in the San1 and Slx5 E3s are localized in intrinsically disordered regions (IDRs) [ 17 , 18 ]. Therefore, we next analyzed the top 200 human lysine desert proteins for intrinsic disorder within the lysine deserts, noting that lysines in general are modestly enriched in IDRs [ 28 , 29 ]. Using the MobiDB database [ 30 ] annotation of intrinsic disorder, the average disorder for proteins in the human proteome was 26%, or 0.26 ± 0.23 (mean ± standard deviation).…”
Section: Resultsmentioning
confidence: 99%
“…Indeed, cysteine is rarely found in RAD23A and UBQLN1 orthologues, which we speculate is because cysteine is generally rare in IDRs [ 33 ]. Conversely, we note that lysine is common in IDRs [ 28 , 29 ]. Fig.…”
In terms of its relative frequency, lysine is a common amino acid in the human proteome. However, by bioinformatics we find hundreds of proteins that contain long and evolutionarily conserved stretches completely devoid of lysine residues. These so-called lysine deserts show a high prevalence in intrinsically disordered proteins with known or predicted functions within the ubiquitin-proteasome system (UPS), including many E3 ubiquitin-protein ligases and UBL domain proteasome substrate shuttles, such as BAG6, RAD23A, UBQLN1 and UBQLN2. We show that introduction of lysine residues into the deserts leads to a striking increase in ubiquitylation of some of these proteins. In case of BAG6, we show that ubiquitylation is catalyzed by the E3 RNF126, while RAD23A is ubiquitylated by E6AP. Despite the elevated ubiquitylation, mutant RAD23A appears stable, but displays a partial loss of function phenotype in fission yeast. In case of UBQLN1 and BAG6, introducing lysine leads to a reduced abundance due to proteasomal degradation of the proteins. For UBQLN1 we show that arginine residues within the lysine depleted region are critical for its ability to form cytosolic speckles/inclusions. We propose that selective pressure to avoid lysine residues may be a common evolutionary mechanism to prevent unwarranted ubiquitylation and/or perhaps other lysine post-translational modifications. This may be particularly relevant for UPS components as they closely and frequently encounter the ubiquitylation machinery and are thus more susceptible to nonspecific ubiquitylation.
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