1H, 15N and 13C assignments of the dimeric ribosome binding domain of trigger factor from Escherichia coli

Trigger factor (TF) is a multi-domain molecular chaperone that binds to the bacterial ribosome at the tunnel exit from which nascent polypeptides emerge. We present here the NMR assignments of the ribosome binding domain (RBD) of TF from Escherichia coli as a stable 26 kDa dimer, using conditions that are similar to a crystallographic study from which an X-ray crystal structure of the identical construct was determined.     

1H, 15N and 13C assignments of the dimeric C-terminal domain of HIV-1 capsid protein

HIV-1 capsid protein (CA) encloses the viral RNA genome and forms a conical-shaped particle in the mature HIV-1 virion, with orderly capsid assembly and disassembly critically important for viral infectivity. The 231 residue CA is composed of two helical domains, connected by a short linker sequence. In solution, CA exhibits concentration dependent dimerization which is mediated by the C-terminal domain (CTD). Here, we present nearly complete ¹H, ¹⁵N and ¹³C assignments for the 20 kDa homodimeric CA–CTD, a prerequisite for structural characterization of the CA–CTD dimer.     

1H, 13C, and 15N NMR assignments of the actinoporin Sticholysin I

Sticholysin I is an actinoporin, a pore forming toxin, of 176 aminoacids produced by the sea anemone Stichodactyla heliantus. Isotopically labelled ¹³C/¹⁵N recombinant protein was produced in E. coli. Here we report the complete NMR ¹⁵N, ¹³C and ¹H chemical shifts assignments of Stn I at pH 4.0 and 25°C (BMRB No. 15927).     

1H, 13C and 15N resonance assignments of rhodanese GlpE from Escherichia coli

Rhodanese catalyzes the sulfur-transfer reaction in which a sulfur atom is transferred from thiosulfate to cyanide by a double-displacement mechanism. During the reaction, a persulfide-intermediate form of rhodanese is generated by the reaction of a conserved active cysteine residue with thiosulfate. Escherichia coli GlpE is a prototype for the single-domain rhodanese superfamily. Though there are some studies on rhodaneses, the molecular mechanism of the catalytic activity of rhodaneses is still unclear. Herein, we report the resonance assignments of (1)H, (13)C and (15)N atoms of E. coli GlpE, which provides the basis for further structural, dynamic and functional studies of rhodaneses using NMR technique.     

1H, 13C and 15N NMR assignments of the Escherichia coli Orf135 protein

Escherichia coli Orf135 protein is thought to be an enzyme that efficiently hydrolyzes oxidatively damaged nucleotides such as 2-hydroxy-dATP, 8-hydroxy-dGTP and 5-hydroxy-CTP, in addition to 5-methyl-dCTP, dCTP and CTP, thus preventing mutations in cells caused by unfavorable base pairing. Nucleotide pool sanitization by Orf135 is important since organisms are continually subjected to potential damage by reactive oxygen species produced during respiration. It is known that the frequency of spontaneous and H₂O₂-induced mutations is two to threefold higher in the orf135 ^- strain compared with the wild-type. Orf135 is a member of the Nudix family of proteins which hydrolyze nucleoside diphosphate derivatives. Nudix hydrolases are characterized by the presence of a conserved motif, although they recognize various substrates and possess a variety of substrate binding pockets. We are interested in delineating the mechanism by which Orf135 recognizes oxidatively damaged nucleotides. To this end, we are investigating the tertiary structure of Orf135 and its interaction with substrate using NMR. Herein, we report on the ¹H, ¹³C and ¹⁵N resonance assignments of Orf135, which should contribute towards a structural understanding of Orf135 and its interaction with substrate.     

1H, 13C and 15N resonance assignments of RNA pyrophosphohydrolase RppH from Escherichia coli

The mRNA degradation is an important regulatory mechanism which controls gene expression by limiting the number of translation times. Previous studies demonstrated that this process is essential for organisms. Escherichia coli RNA pyrophosphohydrolase (RppH) is an enzyme that triggers mRNA degradation by removing the 5′ pyrophosphate, which is a rate-determining step. In order to understand the molecular mechanism of the biological function, the structural information of RppH is required. Herein, we report the resonance assignments of ¹H, ¹⁵N, ¹³C atoms of the E. coli RppH.     

1H, 15N and 13C assignments of yellow fluorescent protein (YFP) Venus

We present here the backbone and side-chain NMR assignments of YFP Venus, a 238-residue protein that emits yellow fluorescence in its native state. Venus is a variant of the green fluorescent protein (GFP), which has improved chromophore maturation and brightness, and the photochemistry and photophysics of which are insensitive to experimental conditions, such as the pH value and buffer content, making it a favourable biomarker.     

1H, 13C and 15N resonance assignments of Rpn9, a regulatory subunit of 26S proteasome from Saccharomyces cerevisiae

The 26S proteasome is an essential molecular machine for specific protein degradation in eukaryotic cells. The 26S proteasome is formed by a central 20S core particle capped by two 19S regulatory particle (RP) at both ends. The Rpn9 protein is a non-ATPase subunit located in the lid complex of the 19S RP, and is identified to be essential for efficient assembly of yeast 26S proteasome. Bioinformatics analysis of Saccharomyces cerevisiae Rpn9 suggested it contains a PCI domain at the C-terminal region. However, high-resolution structures of either the PCI domain or the full-length Rpn9 still remain elusive. Herein, we report the chemical shift assignments of ¹H, ¹³C and ¹⁵N atoms of the individual N- and C-domains, as well as full-length S. cerevisiae Rpn9, which provide the basis for further structural and functional studies of Rpn9 using solution NMR technique.     

1H, 15N and 13C resonance assignments and secondary structure determination of the RNA-binding domain of E. coli rho protein

Summary Protein fragments containing the RNA-binding domain of Escherichia coli rho protein have been over-expressed in E. coli. NMR spectra of the fragment containing residues 1–116 of rho protein (Rho116) show that a region of this protein is unfolded in solution. Addition of (dC)₁₀ to this fragment stabilizes the folded form of the protein. The fragment comprising residues 1–130 of rho protein (Rho130) is found to be stably folded, both in the absence and presence of nucleic acid. NMR studies of the complex of Rho 130 with RNA and DNA oligonucleotides indicate that the binding-site size, affinity, and specificity of Rho 130 are similar to those of intact rho protein; therefore, Rho 130 is a suitable model of the RNA-binding domain of rho protein. NMR line widths as well as titration experiments of Rho130 complexed with oligonucleotides of various lengths suggest that Rho130 forms oligomers in the presence of longer oligonucleotides. ¹H, ¹⁵N and ¹³C resonance assignments were facilitated by the utilization of two pulse sequences, CN-NOESY and CCH-TOCSY. The secondary structure of unliganded Rho130 has been determined by NMR techniques, and it is clear that the RNA-binding domain of rho is more structurally similar to the cold shock domain than to the RNA recognition motif.Abbreviations Rho116, Rho130 protein containing the first 116 (130) residues of rho - CSD cold shock domain - RRM RNA recognition motif - RBD RNA-binding domain - IPTG isopropyl -D-thiogalactopyranoside - EDTA ethylenediaminetetraacetic acid - NOE nuclear Overhauser enhancement     

1H, 15N, and 13C chemical shift assignments of calcium-binding protein 1 (CaBP1)

Calcium-binding protein 1 (CaBP1) regulates inositol 1,4,5-trisphosphate receptors (InsP₃Rs) and a variety of voltage-gated Ca²⁺ channels in the brain. We report complete NMR chemical shift assignments of Ca²⁺-free CaBP1 (residues 1–167, BMRB no. 15197).     

1H, 13C, and 15N resonance assignments of human phosphohistidine phosphatase 1 (PHPT1)

Phosphohistidine phosphatase 1 (PHPT1) is the first protein histidine phosphatase identified in vertebrates. The NMR assignments of human PHPT1 are essential for solution structure determination and NMR study of the protein interactions of PHPT1 with its potential substrates.     

1H, 13C and 15N resonance assignments of the rhodanese domain of YgaP from Escherichia coli

Rhodanese domain is a ubiquitous structural module commonly found in bacterial, archaeal and eukaryotic cells. Growing evidence indicates that rhodanese domains act as the carrier of reactive sulfur atoms by forming persulfide intermediates in distinct metabolic pathways. YgaP, a membrane protein consisting of a rhodanese domain and a C-terminal transmembrane segment, is the only membrane-associated rhodanese in Escherichia coli. Herein, we report the resonance assignments of ¹H, ¹³C and ¹⁵N atoms of rhodanese domain of YgaP. Totally, chemical shifts of more than 95% of the atoms were assigned.     

1H, 15N and 13C assignments of the Rhipicephalus (Boophilus) microplus anti-microbial peptide microplusin

Microplusin, a Rhipicephalus (Boophilus) microplus anti-microbial peptide (AMP) is the first member of a new family of cysteine-rich AMPs with histidine-rich regions at the N- and C-termini, which is being fully characterized by biophysical and biochemical methods. Here we report the NMR resonance assignments for ¹H, ¹⁵N, and ¹³C nuclei in the backbone and side chains of the microplusin as basis for further studies of structure, backbone dynamics and interactions mapping.