Biomedical subjects

Qingqiu Huang

Publications and source records attributed to Qingqiu Huang.

15 recordsLinked to original sources

PubMed2006

Structure of human MRG15 chromo domain and its binding to Lys36-methylated histone H3.

Human MRG15 is a transcription factor that plays a vital role in embryonic development, cell proliferation and cellular senescence. It comprises a putative chromo domain in the N-terminal part that has been shown to participate in chromatin remodeling and transcription regulation. We report here the crystal structure of human MRG15 chromo domain at 2.2 A resolution. The MRG15 chromo domain consists of a beta-barrel and a long alpha-helix and assumes a structure more similar to the Drosophila MOF chromo barrel domain than the typical HP1/Pc chromo domains. The beta-barrel core contains a hydrophobic pocket formed by three conserved aromatic residues Tyr26, Tyr46 and Trp49 as a potential binding site for a modified residue of histone tail. However, the binding groove for the histone tail seen in the HP1/Pc chromo domains is pre-occupied by an extra beta-strand. In vitro binding assay results indicate that the MRG15 chromo domain can bind to methylated Lys36, but not methylated Lys4, Lys9 and Lys27 of histone H3. These data together suggest that the MRG15 chromo domain may function as an adaptor module which can bind to a modified histone H3 in a mode different from that of the HP1/Pc chromo domains.

Amino Acid Sequence↗

PubMed2006

Crystal structure of mammalian cysteine dioxygenase. A novel mononuclear iron center for cysteine thiol oxidation.

Cysteine dioxygenase is a mononuclear iron-dependent enzyme responsible for the oxidation of cysteine with molecular oxygen to form cysteine sulfinate. This reaction commits cysteine to either catabolism to sulfate and pyruvate or the taurine biosynthetic pathway. Cysteine dioxygenase is a member of the cupin superfamily of proteins. The crystal structure of recombinant rat cysteine dioxygenase has been determined to 1.5-A resolution, and these results confirm the canonical cupin beta-sandwich fold and the rare cysteinyltyrosine intramolecular cross-link (between Cys(93) and Tyr(157)) seen in the recently reported murine cysteine dioxygenase structure. In contrast to the catalytically inactive mononuclear Ni(II) metallocenter present in the murine structure, crystallization of a catalytically competent preparation of rat cysteine dioxygenase revealed a novel tetrahedrally coordinated mononuclear iron center involving three histidines (His(86), His(88), and His(140)) and a water molecule. Attempts to acquire a structure with bound ligand using either cocrystallization or soaking crystals with cysteine revealed the formation of a mixed disulfide involving Cys(164) near the active site, which may explain previously observed substrate inhibition. This work provides a framework for understanding the molecular mechanisms involved in thiol dioxygenation and sets the stage for exploration of the chemistry of both the novel mononuclear iron center and the catalytic role of the cysteinyl-tyrosine linkage.

Amino Acid Sequence↗

PubMed2005

Blocking effect and crystal structure of natrin toxin, a cysteine-rich secretory protein from Naja atra venom that targets the BKCa channel.

Cysteine-rich secretory proteins (CRISPs) are widespread in snake venoms. Some members of these CRISPs recently have been found to block L-type Ca(2+) channels or cyclic nucleotide-gated ion (CNG) channels. Here, natrin purified from Naja atra venom, a member of the CRISP family, can induce a further contractile response in the endothelium-denuded thoracic aorta of mouse which has been contracted by a high-K(+) solution. Further experiments show it can block the high-conductance calcium-activated potassium (BK(Ca)) channel in a concentration-dependent manner with an IC(50) of 34.4 nM and a Hill coefficient of 1.02, which suggests that only a single natrin molecule is required to bind an ion channel to block BK(Ca) current. The crystal structure of natrin displaying two domains in tandem shows its cysteine-rich domain (CRD) has relatively independent flexibility, especially for the C-terminal long loop (loop I) of CRD to participate in the interface of two domains. On the basis of previous studies of CNG channel and L-Ca(2+) channel blockers, and the sequence and structural comparison of natrin and stecrisp, the deviation of the vital loop I of CRD is suggested to contribute to different effects of some CRISPs in protein-protein interaction.

Amino Acid Sequence↗

PubMed2005

Crystal structures of Fms1 and its complex with spermine reveal substrate specificity.

Fms1 is a rate-limiting enzyme for the biosynthesis of pantothenic acid in yeast. Fms1 has polyamine oxidase (PAO) activity, which converts spermine into spermidine and 3-aminopropanal. The 3-aminopropanal is further oxidized to produce beta-alanine, which is necessary for the biosynthesis of pantothenic acid. The crystal structures of Fms1 and its complex with the substrate spermine have been determined using the single-wavelength anomalous diffraction (SAD) phasing method. Fms1 consists of an FAD-binding domain, with Rossmann fold topology, and a substrate-binding domain. The active site is a tunnel located at the interface of the two domains. The substrate spermine binds to the active site mainly via hydrogen bonds and hydrophobic interactions. In the complex, C11 but not C9 of spermine is close enough to the catalytic site (N5 of FAD) to be oxidized. Therefore, the products are spermidine and 3-aminopropanal, rather than 3-(aminopropyl) 4-aminobutyraldehyde and 1,3-diaminoprone.

Amino Acid Sequence↗

PubMed2004

Crystal structure of the cysteine-rich secretory protein stecrisp reveals that the cysteine-rich domain has a K+ channel inhibitor-like fold.

Stecrisp from Trimeresurus stejnegeri snake venom belongs to a family of cysteine-rich secretory proteins (CRISP) that have various functions related to sperm-egg fusion, innate host defense, and the blockage of ion channels. Here we present the crystal structure of stecrisp refined to 1.6-angstrom resolution. It shows that stecrisp contains three regions, namely a PR-1 (pathogenesis-related proteins of group1) domain, a hinge, and a cysteine-rich domain (CRD). A conformation of solvent-exposed and -conserved residues (His60, Glu75, Glu96, and His115) in the PR-1 domain similar to that of their counterparts in homologous structures suggests they may share some molecular mechanism. Three flexible loops of hypervariable sequence surrounding the possible substrate binding site in the PR-1 domain show an evident difference in homologous structures, implying that a great diversity of species- and substrate-specific interactions may be involved in recognition and catalysis. The hinge is fixed by two crossed disulfide bonds formed by four of ten characteristic cysteines in the carboxyl-terminal region and is important for stabilizing the N-terminal PR-1 domain. Spatially separated from the PR-1 domain, CRD possesses a similar fold with two K+ channel inhibitors (Bgk and Shk). Several candidates for the possible functional sites of ion channel blocking are located in a solvent-exposed loop in the CRD. The structure of stecrisp will provide a prototypic architecture for a structural and functional exploration of the diverse members of the CRISP family.

Amino Acid Sequence↗

PubMed2004

Crystallization and preliminary X-ray crystallographic analysis of agkicetin-C from Deinagkistrodon acutus venom.

The crystallization and preliminary crystallographic analysis of agkicetin-C, a well known platelet glycoprotein Ib (GPIb) antagonist from the venom of Deinagkistrodon acutus found in Anhui Province, China is reported. Crystals of agkicetin-C suitable for structure determination were obtained from 1.8 M ammonium sulfate, 40 mM MES pH 6.5 with 2%(v/v) PEG 400. Interestingly, low buffer concentrations of MES seem to be necessary for crystal growth. The crystals of agkicetin-C belong to space group C2, with unit-cell parameters a = 177.5, b = 97.7, c = 106.8 A, beta = 118.5 degrees, and diffract to 2.4 A resolution. Solution of the phase problem by the molecular-replacement method shows that there are four agkicetin-C molecules in the asymmetric unit, with a VM value of 3.4 A3 Da(-1), which corresponds to a high solvent content of approximately 64%. Self-rotation function calculations show a single well defined non-crystallographic twofold axis with features that may represent additional elements of non-crystallographic symmetry.

Agkistrodon↗

PubMed2004

Purification, characterization, crystallization and preliminary X-ray crystallographic analysis of two novel C-type lectin-like proteins: Aall-A and Aall-B from Deinagkistrodon acutus venom.

Aall-A and Aall-B, two novel heterodimeric snake-venom C-type lectin-like proteins (sv-CLPs), were purified from the venom of Deinagkistrodon acutus from Anhui, China. Strikingly, both these proteins can localize on and congregate human erythrocytes, instead of aiming at the common targets of sv-CLPs such as platelet glycoproteins, von Willebrand factors, coagulant factors etc. The crystals of Aall-A belong to space group P2, with unit-cell parameters a = 105.2, b = 56.2, c = 108.7 A, beta = 100.5 degrees , and diffract to 2.0 A resolution, while the crystals of Aall-B belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 36.8, b = 56.5, c = 149.2 A, and diffract to 2.2 A resolution. To our knowledge, this is the first report of sv-CLPs with this unique function and of their preliminary crystallographic analysis.

Amino Acid Sequence↗

PubMed2004

The atomic resolution crystal structure of atratoxin determined by single wavelength anomalous diffraction phasing.

By using single wavelength anomalous diffraction phasing based on the anomalous signal from copper atoms, the crystal structure of atratoxin was determined at the resolution of 1.5 A and was refined to an ultrahigh resolution of 0.87 A. The ultrahigh resolution electron density maps allowed the modeling of 38 amino acid residues in alternate conformations and the location of 322 of 870 possible hydrogen atoms. To get accurate information at the atomic level, atratoxin-b (an analog of atratoxin with reduced toxicity) was also refined to an atomic resolution of 0.92 A. By the sequence and structural comparison of these two atratoxins, Arg(33) and Arg(36) were identified to be critical to their varied toxicity. The effect of copper ions on the distribution of hydrogen atoms in atratoxin was discussed, and the interactions between copper ions and protein residues were analyzed based on a statistical method, revealing a novel pentahedral copper-binding motif.

Amino Acid Motifs↗

PubMed2004

Purification, partial characterization, crystallization and preliminary X-ray diffraction of two cysteine-rich secretory proteins from Naja atra and Trimeresurus stejnegeri venoms.

Cysteine-rich secretory proteins (CRISPs) are widely distributed in mammals and snake venoms. They possess apparent homology but varying functions. The structure of CRISPs has remained elusive. Two novel members of the family, natrin and stecrisp, have been purified from Naja atra and Trimeresurus stejnegeri venoms, respectively. Their crystals diffract X-rays to resolution limits of 2.1 and 1.6 angstroms, respectively, and belong to the orthorhombic system with different space groups, unit-cell parameters and numbers of molecules per asymmetric unit. Their structures will contribute a structural basis for further functional studies of this family.

Amino Acid Sequence↗

PubMed2004

Purification, partial characterization, crystallization and structural determination of AHP-LAAO, a novel L-amino-acid oxidase with cell apoptosis-inducing activity from Agkistrodon halys pallas venom.

A snake-venom protein named AHP-LAAO has been purified from Agkistrodon halys pallas venom using four-stage chromatography. AHP-LAAO is a novel member of the snake-venom L-amino-acid oxidase family. Its amino-acid sequence shows high homology to other members of this family. For L-leucine, the values of k(cat) and K(M) are 31.1 s(-1) and 0.25 mM, respectively. The molecular weight of AHP-LAAO is about 60.7 kDa as determined by MALDI-TOF mass spectrometry. AHP-LAAO can also induce apoptosis of cultured Hela cells. Two sets of diffraction data with similar resolution limits (about 2.5 A) were collected independently at MacCHESS (Cornell High Energy Synchrotron Source, USA) and IHEP (Institute of High Energy Physics, Beijing, China). The crystals belong to space group I2(1)3, with unit-cell parameter a = 169.31 A, corresponding to one molecule in the asymmetric unit and a volume-to-weight ratio of 3.33 A(3) Da(-1). The final structural model is similar to that of L-amino-acid oxidase from Calloselasma rhodostoma venom.

Amino Acid Oxidoreductases↗

PubMed2004

Crystallographic snapshots of Aspergillus fumigatus phytase, revealing its enzymatic dynamics.

Understanding of the atomic movements involved in an enzymatic reaction needs structural information on the active and inactive native enzyme molecules and on the enzyme-substrate, enzyme-intermediate, and enzyme-product(s) complexes. By using the X-ray crystallographic method, four crystal structures of Aspergillus fumigatus phytase were obtained at resolution higher than 1.7 A. The pH-dependent catalytic activity of A. fumigatus phytase was linked to three water molecules that may prevent the substrate from binding and thus block nucleophilic attack of the catalytic imidazole nitrogen. Comparison of various structures also identified the water molecule that attacks the phosphamide bond during the hydrolysis process, and established the hydrolysis pathway of the intermediate. Additionally, two reaction product phosphates were observed at the active site, suggesting a possible product release pathway after hydrolysis of the intermediate. These results can help explain the catalytic mechanism throughout the whole acid phosphatase family, as all key residues are conserved.

6-Phytase↗

PubMed2003

Purification, partial characterization and crystallization of acucetin, a protein containing both disintegrin-like and cysteine-rich domains released by auto-proteolysis of a P-III-type metalloproteinase AaH-IV from Agkistrodon acutus venom.

AaH-IV, a P-III-type metalloproteinase found in Agkistrodon acutus venom, readily cleaves itself to release a stable protein named acucetin at 310 K under neutral and weakly alkaline conditions. A partial amino-acid residue sequence of acucetin indicates that the protein has a high homology to snake-venom proteins containing both disintegrin-like and cysteine-rich domains. Acucetin has been crystallized in space group R32, with hexagonal unit-cell parameters a = b = 155.98, c = 76.07 A. The V(M) value of about 2.97 A(3) Da(-1) suggests the presence of only one molecule in the asymmetric unit.

Agkistrodon↗

PubMed2003

The crystal structure of a novel, inactive, lysine 49 PLA2 from Agkistrodon acutus venom: an ultrahigh resolution, AB initio structure determination.

The crystal structure of acutohaemolysin, a lysine 49 phospholipase A2 protein with 1010 non-hydrogen protein atoms and 232 water molecules, has been determined ab initio using the program SnB at an ultrahigh resolution of 0.8 A. The lack of catalytic activity appears to be related to the presence of Phe102, which prevents the access of substrate to the active site. The substitution of tryptophan for leucine at residue 10 interferes with dimer formation and may be responsible for the additional loss of hemolytic activity. The ultrahigh resolution of the experimental diffraction data permits alternative conformations to be modeled for disordered residues, many hydrogen atoms to be located, the protonation of the Nepsilon2 atom in the catalytic residue His48 to be observed experimentally, and the density of the bonding electrons to be analyzed in detail.

Agkistrodon↗

PubMed2002

Purification, N-terminal sequencing, crystallization and preliminary X-ray diffraction analysis of atratoxin, a new short-chain alpha-neurotoxin from the venom of Naja naja atra.

Atratoxin, a new alpha-neurotoxin purified to homogeneity by a series of liquid chromatographies from the venom of Naja naja atra (mainland Chinese cobra), is a small single-polypeptide alkaline protein with a pI of about 9.5 and molecular weight of 6952 Da estimated by mass spectrometry. Although the sequencing of the N-terminal 15 residues (LECHNQQTTQQPEGG) shows that this neurotoxic protein contains most of the residues, especially at the conserved positions, of the consensus sequence of short-chain alpha-neurotoxins, the natural mutations in the N-terminal Loop-1 presented by the sequence alignment may have structural or functional implications for the interactions between alpha-neurotoxins and related receptors. Single crystals of atratoxin have been grown from drops containing the necessary Cu(2+) ions by the conventional hanging-drop vapour-diffusion method. The crystals diffract X-rays to 1.6 A resolution and belong to space group C222(1), with unit-cell parameters a = 47.36, b = 47.83, c = 91.31 A, corresponding to a volume-to-mass ratio of 1.85 A(3) Da(-1) and two molecules in each asymmetric unit.

Amino Acid Sequence↗

PubMed2002

Purification, crystallization and preliminary X-ray crystallographic analysis of agkaggregin, a C-type lectin-like protein from Agkistrodon acutus venom.

A snake-venom C-type lectin-like protein, agkaggregin, has been isolated from Agkistrodon acutus venom. Agkaggregin has an apparent molecular mass of about 28 kDa and consists of two different types of subunits, an alpha-subunit (approximately 15 kDa) and a beta-subunit (approximately 14 kDa). Agkaggregin has the ability to induce platelet aggregation at concentrations of the order of nanomoles. The agkaggregin crystals grew for nearly a year by hanging-drop vapour diffusion and belong to the I222 space group, with unit-cell parameters a = 64.75, b = 74.21, c = 133.24 A. One asymmetric unit contains one alphabeta heterodimer, corresponding to a volume-to-mass ratio of 2.795 A(3) Da(-1). Agkaggregin may exist in two association forms: an alphabeta heterodimer and a dimer of alphabeta heterodimers that associates during the long process of crystallization.

Crotalid Venoms↗