scholarly journals di-Cysteine Residues of the Arabidopsis thaliana HMA4 C-Terminus Are Only Partially Required for Cadmium Transport

2020 ◽  
Vol 11 ◽  
Author(s):  
Stanislaus Antony Ceasar ◽  
Gilles Lekeux ◽  
Patrick Motte ◽  
Zhiguang Xiao ◽  
Moreno Galleni ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Cysteine Residues ◽  
C Terminus ◽  
Cadmium Transport

scholarly journals Primary- and secondary-structural analysis of a unique prokaryotic metallothionein from a Synechococcus sp. cyanobacterium

1988 ◽  
Vol 251 (3) ◽  
pp. 691-699 ◽  
Author(s):  
R W Olafson ◽  
W D McCubbin ◽  
C M Kay
Keyword(s):  
Amino Acid ◽  
Metal Binding ◽  
Helical Structure ◽  
Basic Amino Acid ◽  
Cysteine Residues ◽  
Amino Acid Residues ◽  
C Terminus ◽  
Empirical Relations ◽  
Heavy Metal Binding ◽  
Synechococcus Sp

Biochemical and physiological studies of Synechococcus cyanobacteria have indicated the presence of a low-Mr heavy-metal-binding protein with marked similarity to eukaryotic metallothioneins (MTs). We report here the characterization of a Synechococcus prokaryotic MT isolated by gel-permeation and reverse-phase chromatography. The large number of variants of this molecule found during chromatographic separation could not be attributed to the presence of major isoproteins as assessed by amino acid analysis and amino acid sequencing of isoforms. Two of the latter were shown to have identical primary structures that differed substantially from the well-described eukaryotic MTs. In addition to six long-chain aliphatic residues, two aromatic residues were found adjacent to one another near the centre of the molecule, making this the most hydrophobic MT to be described. Other unusual features included a pair of histidine residues located in repeating Gly-His-Thr-Gly sequences near the C-terminus and a complete lack of association of hydroxylated residues with cysteine residues, as is commonly found in eukaryotes. Similarly, aside from a single lysine residue, no basic amino acid residues were found adjacent to cysteine residues in the sequence. Most importantly, sequence alignment analyses with mammalian, invertebrate and fungal MT sequences showed no statistically significant homology aside from the presence of Cys-Xaa-Cys structures common to all MTs. On the other hand, like other MTs, the prokaryotic molecule appears to be free of alpha-helical structure but has a considerable amount of beta-structure, as predicted by both c.d. measurements and the Chou & Fasman empirical relations. Considered together, these data suggested that some similarity between the metal-thiolate clusters of the prokaryote and eukaryote MTs may exist.

scholarly journals Comparison of two Turnip mosaic virus P1 proteins in their ability to co-localize with the Arabidopsis thaliana G3BP-2 protein

Virus Genes ◽  
2021 ◽  
Vol 57 (2) ◽  
pp. 233-237
Author(s):  
Hendrik Reuper ◽  
Björn Krenz
Keyword(s):  
Arabidopsis Thaliana ◽  
Mosaic Virus ◽  
Specific Interaction ◽  
Viral Proteins ◽  
Turnip Mosaic Virus ◽  
Stress Conditions ◽  
C Terminus ◽  
Positive Sense ◽  
Key Factor ◽  
Large Host

AbstractTurnip mosaic virus (TuMV), belonging to the genus Potyvirus (family Potyviridae), has a large host range and consists of a single-stranded positive sense RNA genome encoding 12 proteins, including the P1 protease. This protein which is separated from the polyprotein by cis cleavage at its respective C-terminus, has been attributed with different functions during potyviral infection of plants. P1 of Turnip mosaic virus (P1-TuMV) harbors an FGSF-motif and FGSL-motif at its N-terminus. This motif is predicted to be a binding site for the host Ras GTPase-activating protein-binding protein (G3BP), which is a key factor for stress granule (SG) formation in mammalian systems and often targeted by viruses to inhibit SG formation. We therefore hypothesized that P1-TuMV might interact with G3BP to control and regulate plant SGs to optimize cellular conditions for the production of viral proteins. Here, we analyzed the co-localization of the Arabidopsis thaliana G3BP-2 with the P1 of two TuMV isolates, namely UK 1 and DEU 2. Surprisingly, P1-TuMV-DEU 2 co-localized with AtG3BP-2 under abiotic stress conditions, whereas P1-TuMV-UK 1 did not. AtG3BP-2::RFP showed strong SGs formation after stress, while P1-UK 1::eGFP maintained a chloroplastic signal under stress conditions, the signal of P1-DEU 2::eGFP co-localized with that of AtG3BP-2::RFP. This indicates a specific interaction between P1-DEU 2 and the AtG3BP family which is not solely based on the canonical interaction motifs.

IRON MAN interacts with BRUTUS to maintain iron homeostasis in Arabidopsis

2021 ◽  
Vol 118 (39) ◽  
pp. e2109063118
Author(s):  
Yang Li ◽  
Cheng Kai Lu ◽  
Chen Yang Li ◽  
Ri Hua Lei ◽  
Meng Na Pu ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Transcription Factors ◽  
Iron Homeostasis ◽  
Fe Deficiency ◽  
Small Peptides ◽  
Interaction Domain ◽  
C Terminus ◽  
Positive Regulators ◽  
Fe Homeostasis ◽  
And Function

IRON MAN (IMA) peptides, a family of small peptides, control iron (Fe) transport in plants, but their roles in Fe signaling remain unclear. BRUTUS (BTS) is a potential Fe sensor that negatively regulates Fe homeostasis by promoting the ubiquitin-mediated degradation of bHLH105 and bHLH115, two positive regulators of the Fe deficiency response. Here, we show that IMA peptides interact with BTS. The C-terminal parts of IMA peptides contain a conserved BTS interaction domain (BID) that is responsible for their interaction with the C terminus of BTS. Arabidopsis thaliana plants constitutively expressing IMA genes phenocopy the bts-2 mutant. Moreover, IMA peptides are ubiquitinated and degraded by BTS. bHLH105 and bHLH115 also share a BID, which accounts for their interaction with BTS. IMA peptides compete with bHLH105/bHLH115 for interaction with BTS, thereby inhibiting the degradation of these transcription factors by BTS. Genetic analyses suggest that bHLH105/bHLH115 and IMA3 have additive roles and function downstream of BTS. Moreover, the transcription of both BTS and IMA3 is activated directly by bHLH105 and bHLH115 under Fe-deficient conditions. Our findings provide a conceptual framework for understanding the regulation of Fe homeostasis: IMA peptides protect bHLH105/bHLH115 from degradation by sequestering BTS, thereby activating the Fe deficiency response.

scholarly journals In-Situ Flexibility of Both Redox-States of the Chloroplast Regulatory Protein CP12

2021 ◽  
Author(s):  
Helene Launay ◽  
Hui Shao ◽  
Olivier Bornet ◽  
Francois-Xavier Cantrelle ◽  
Regine Lebrun ◽  
...  
Keyword(s):  
Regulatory Protein ◽  
Cell Extract ◽  
Proton Exchange ◽  
Conformational Exchange ◽  
Amide Proton ◽  
Cysteine Residues ◽  
Intrinsically Disordered ◽  
Amide Proton Exchange ◽  
C Terminus ◽  
Helical Turn

In the chloroplast, Calvin-Benson-Bassham enzymes are active in the reducing environment imposed in the light via the electrons from the photosystems. In the dark these enzymes are inhibited, and this regulation is mainly mediated via oxidation of key regulatory cysteine residues. CP12 is a small protein that plays a role in this regulation with four cysteine residues that undergo a redox transition. Using amide-proton exchange with solvent measured by nuclear magnetic resonance (NMR) and mass-spectrometry, we confirmed that reduced CP12 is intrinsically disordered. Using real-time NMR, we showed that the oxidation of the two disulfide bridges are simultaneous. In oxidized CP12, the C23-C31 pair is in a region that undergoes a conformational exchange in the NMR-intermediate timescale. The C66-C75 pair is in the C-terminus that folds into a stable helical turn. We confirmed that these structural states exist in a physiologically relevant environment that is, in cell extract from Chlamydomonas reinhardtii. Consistent with these structural equilibria, the reduction is slower for the C66-C75 pair compared to the C23-C31 pair. Finally, the redox mid-potentials for the two cysteine pairs differ and are similar to those found for phosphoribulokinase and glyceraldehyde 3-phosphate dehydrogenase, that we relate to the regulatory role of CP12.

Profiling sulfenylated cysteine residues in Arabidopsis thaliana under oxidative stress

2012 ◽  
Vol 53 ◽  
pp. S224-S225
Author(s):  
M.S. Akter⁎ ◽  
K. Wahni ◽  
C. Waszczak ◽  
S. Carpentier ◽  
F.V. Breusegem ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Arabidopsis Thaliana ◽  
Cysteine Residues

scholarly journals Engineering the nucleotide coenzyme specificity and sulfhydryl redox sensitivity of two stress-responsive aldehyde dehydrogenase isoenzymes of Arabidopsis thaliana

2011 ◽  
Vol 434 (3) ◽  
pp. 459-471 ◽  
Author(s):  
Naim Stiti ◽  
Isaac O. Adewale ◽  
Jan Petersen ◽  
Dorothea Bartels ◽  
Hans-Hubert Kirch
Keyword(s):  
Arabidopsis Thaliana ◽  
Protein Structures ◽  
Aromatic Aldehydes ◽  
Carbon Chain ◽  
Cysteine Residues ◽  
Carbon Chain Length ◽  
Coenzyme Specificity ◽  
Coenzyme Binding ◽  
Catalytic Cysteine ◽  
Binding Cleft

Lipid peroxidation is one of the consequences of environmental stress in plants and leads to the accumulation of highly toxic, reactive aldehydes. One of the processes to detoxify these aldehydes is their oxidation into carboxylic acids catalyzed by NAD(P)+-dependent ALDHs (aldehyde dehydrogenases). We investigated kinetic parameters of two Arabidopsis thaliana family 3 ALDHs, the cytosolic ALDH3H1 and the chloroplastic isoform ALDH3I1. Both enzymes had similar substrate specificity and oxidized saturated aliphatic aldehydes. Catalytic efficiencies improved with the increase of carbon chain length. Both enzymes were also able to oxidize α,β-unsaturated aldehydes, but not aromatic aldehydes. Activity of ALDH3H1 was NAD+-dependent, whereas ALDH3I1 was able to use NAD+ and NADP+. An unusual isoleucine residue within the coenzyme-binding cleft was responsible for the NAD+-dependence of ALDH3H1. Engineering the coenzyme-binding environment of ALDH3I1 elucidated the influence of the surrounding amino acids. Enzyme activities of both ALDHs were redox-sensitive. Inhibition was correlated with oxidation of both catalytic and non-catalytic cysteine residues in addition to homodimer formation. Dimerization and inactivation could be reversed by reducing agents. Mutant analysis showed that cysteine residues mediating homodimerization are located in the N-terminal region. Modelling of the protein structures revealed that the redox-sensitive cysteine residues are located at the surfaces of the subunits.

scholarly journals Role of C-Terminal Cysteine Residues of Aspergillus fumigatus Allergen Asp f 4 in Immunoglobulin E Binding

2004 ◽  
Vol 11 (2) ◽  
pp. 261-265 ◽  
Author(s):  
Harikrishnan Ramachandran ◽  
Banani Banerjee ◽  
Paul A. Greenberger ◽  
Kevin J. Kelly ◽  
Jordan N. Fink ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Conformational Changes ◽  
Immunoglobulin E ◽  
Allergic Bronchopulmonary Aspergillosis ◽  
Allergic Diseases ◽  
Cysteine Residues ◽  
Conformational Structure ◽  
C Terminus ◽  
Function Relationship

ABSTRACT Among the several allergens cloned and expressed from Aspergillus fumigatus, Asp f 4 is a major one associated with allergic bronchopulmonary aspergillosis (ABPA). The structure-function relationship of allergens is important in understanding the immunopathogenesis, diagnosis, and treatment of allergic diseases. These include the epitopes, conformational or linear, deletion of the N or C terminus or both N and C termini, and glycosylation or nonglycosylation, all of which affect immune responses. Similarly, the role of cysteine residues present in allergens may yield useful information regarding the conformational structure of allergens and the immunoglobulin E (IgE) epitope interaction. Such information may help in developing new strategies towards immunotherapy. In order to define the role of cysteine in the interaction of the antibody with Asp f 4, we have constructed mutants by selectively deleting cysteine residues from the C-terminal region of the Asp f 4. Immunological evaluation of these engineered recombinant constructs was conducted by using sera from patients with ABPA, Aspergillus skin test-positive asthmatics, and healthy controls. The results demonstrate strong IgE binding with Asp f 4 and two truncated mutants, Asp f 41-234 (amino acids [aa] 1 to 234) and Asp f 41-241 (aa 1 to 241), while another mutant, Asp f 41-196 (aa 1 to 196), showed reactivity with fewer patients. The result suggests that deletion of cysteines and the alteration of IgE epitopes at the C-terminal end resulted in conformational changes, which may have a potential role in the immunomodulation of the disease.

scholarly journals Four Conserved Cysteine Residues of the Hepatitis B Virus Polymerase Are Critical for RNA Pregenome Encapsidation

2009 ◽  
Vol 83 (16) ◽  
pp. 8032-8040 ◽  
Author(s):  
Seahee Kim ◽  
Jehan Lee ◽  
Wang-Shick Ryu
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Reverse Transcriptase ◽  
Rna Binding ◽  
Cysteine Residues ◽  
Zinc Finger Motif ◽  
Terminal Protein ◽  
C Terminus ◽  
B Virus ◽  
Putative Zinc Finger

ABSTRACT Hepadnaviruses replicate via reverse transcription of an RNA template, the pregenomic RNA (pgRNA). Although hepadnaviral polymerase (Pol) and retroviral reverse transcriptase are distantly related, some of their features are distinct. In particular, Pol contains two additional N-terminal subdomains, the terminal protein and spacer subdomains. Since much of the spacer subdomain can be deleted without detrimental effects to hepatitis B virus (HBV) replication, this subdomain was previously thought to serve only as a spacer that links the terminal protein and reverse transcriptase subdomains. Unexpectedly, we found that the C terminus of the spacer subdomain is indispensable for the encapsidation of pgRNA. Alanine-scanning mutagenesis revealed that four conserved cysteine residues, three at the C terminus of the spacer subdomain and one at the N terminus of the reverse transcriptase subdomain, are critical for encapsidation. The inability of the mutant Pol proteins to incorporate into nucleocapsid particles, together with other evidence, argued that the four conserved cysteine residues are critical for RNA binding. One implication is that these four cysteine residues might form a putative zinc finger motif. Based on these findings, we speculate that the RNA binding activity of HBV Pol may be mediated by this newly identified putative zinc finger motif.

scholarly journals Molecular cloning and functional expression of bovine spleen ecto-NAD+ glycohydrolase: structural identity with human CD38

1999 ◽  
Vol 345 (1) ◽  
pp. 43-52 ◽  
Author(s):  
Angélique AUGUSTIN ◽  
Hélène MULLER-STEFFNER ◽  
Francis SCHUBER
Keyword(s):  
Transmembrane Protein ◽  
Functional Expression ◽  
Full Length ◽  
Cysteine Residues ◽  
Sequence Information ◽  
Functional Domain ◽  
High Sequence Identity ◽  
Full Length Cdna ◽  
Nad Glycohydrolase ◽  
C Terminus

Bovine spleen ecto-NAD+ glycohydrolase, an archetypal member of the mammalian membrane-associated NAD(P)+ glycohydrolase enzyme family (EC 3.2.2.6), displays catalytic features similar to those of CD38, i.e. a protein originally described as a lymphocyte differentiation marker involved in the metabolism of cyclic ADP-ribose and signal transduction. Using amino acid sequence information obtained from NAD+ glycohydrolase and from a truncated and hydrosoluble form of the enzyme (hNADase) purified to homogeneity, a full-length cDNA clone was obtained. The deduced sequence indicates a protein of 278 residues with a molecular mass of 31.5 kDa. It predicts that bovine ecto-NAD+ glycohydrolase is a type II transmembrane protein, with a very short intracellular tail. The bulk of the enzyme, which is extracellular and contains two potential N-glycosylation sites, yields the fully catalytically active hNADase which is truncated by 71 residues. Transfection of HeLa cells with the full-length cDNA resulted in the expression of the expected NAD+ glycohydrolase, ADP-ribosyl cyclase and GDP-ribosyl cyclase activities at the surface of the cells. The bovine enzyme, which is the first ‘classical’ NAD(P)+ glycohydrolase whose structure has been established, presents a particularly high sequence identity with CD38, including the presence of 10 strictly conserved cysteine residues in the ectodomain and putative catalytic residues. However, it lacks two otherwise conserved cysteine residues near its C-terminus. Thus hNADase, the truncated protein of 207 amino acids, represents the smallest functional domain endowed with all the catalytic activities of CD38/NAD+ glycohydrolases so far identified. Altogether, our data strongly suggest that the cloned bovine spleen ecto-NAD+ glycohydrolase is the bovine equivalent of CD38.

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