Crystal Structure of Human Leukocyte Cell-derived Chemotaxin 2 (LECT2) Reveals a Mechanistic Basis of Functional Evolution in a Mammalian Protein with an M23 Metalloendopeptidase Fold.

Human leukocyte cell-derived chemotaxin 2 (LECT2), which is predominantly expressed in the liver, is a multifunctional protein. LECT2 is becoming a potential therapeutic target for several diseases of worldwide concern such as rheumatoid arthritis, hepatocellular carcinoma, and obesity. Here, we present the crystal structure of LECT2, the first mammalian protein whose structure contains an M23 metalloendopeptidase fold.

A secreted protein with plant-specific cysteine-rich motif functions as a mannose-binding lectin that exhibits antifungal activity.

Plants have a variety of mechanisms for defending against plant pathogens and tolerating environmental stresses such as drought and high salinity. Ginkbilobin2 (Gnk2) is a seed storage protein in gymnosperm that possesses antifungal activity and a plant-specific cysteine-rich motif (domain of unknown function26 [DUF26]). The Gnk2-homologous sequence is also observed in an extracellular region of cysteine-rich repeat receptor-like kinases that function in response to biotic and abiotic stresses.

Calcium-dependent structural changes in human reticulocalbin-1.

Human reticulocalbin-1 (hRCN1) has six EF-hand motifs and binds Ca(2+). hRCN1 is a member of the CREC family localized in the secretory pathway, and its cellular function remains unclear. In this study, we established a new bacterial expression and purification procedure for hRCN1.

A thermoacidophile-specific protein family, DUF3211, functions as a fatty acid carrier with novel binding mode.

STK_08120 is a member of the thermoacidophile-specific DUF3211 protein family from Sulfolobus tokodaii strain 7. Its molecular function remains obscure, and sequence similarities for obtaining functional remarks are not available. In this study, the crystal structure of STK_08120 was determined at 1.

Crystallization and preliminary X-ray analysis of human leukocyte cell-derived chemotaxin 2 (LECT2).

Human leukocyte cell-derived chemotaxin 2 (LECT2) is a chemotactic factor for neutrophils that plays multifunctional roles in liver regeneration, regulation of neuritic development and proliferation of chondrocytes and osteoblasts. In addition, the C-terminal region of LECT2 belongs to the zinc metalloendopeptidase M23 (PF01551) family. Purified LECT2 was crystallized using the sitting-drop vapour-diffusion method at 293 K.

Expression, high-pressure refolding and purification of human leukocyte cell-derived chemotaxin 2 (LECT2).

Human leukocyte cell-derived chemotaxin 2 (LECT2) is a chemotactic factor for neutrophils and a 16-kDa secreted protein consisting of 133 amino acids with three intramolecular disulfide bonds. Here, we propose an efficient method for the preparation of human LECT2 using a high hydrostatic pressure (HHP, 200 MPa) refolding technique. When LECT2 was over-expressed in Escherichia coli cells, most of LECT2 molecules became insoluble inclusion bodies (IBs).

Structure of the Ca(2+)-saturated C-terminal domain of scallop troponin C in complex with a troponin I fragment.

Troponin C (TnC) is the Ca(2+)-sensing subunit of troponin that triggers the contraction of striated muscles. In scallops, the striated muscles consume little ATP energy in sustaining strong contractile forces. The N-terminal domain of TnC works as the Ca(2+) sensor in vertebrates, whereas scallop TnC uses the C-terminal domain as the Ca(2+) sensor, suggesting that there are differences in the mechanism of the Ca(2+)-dependent regulation of muscles between invertebrates and vertebrates.

Expression, purification, crystallization and preliminary X-ray analysis of carbonyl reductase S1 from Candida magnoliae.

The NADPH-dependent carbonyl reductase S1 from Candida magnoliae stereoselectively catalyzes the reduction of ethyl 4-chloro-3-oxobutanoate (COBE) to ethyl (S)-4-chloro-3-hydroxybutanoate (CHBE), which is a chiral compound valuable as a building block for pharmaceuticals. Carbonyl reductase S1 was expressed in Escherichia coli and purified by Ni-affinity, ion-exchange and size-exclusion chromatography. Crystals of carbonyl reductase S1 were obtained by the sitting-drop vapour-diffusion method using PEG 400 as a precipitant.

Crystallization and preliminary X-ray crystallographic analysis of dioscorin from Dioscorea japonica.

Dioscorin, the major tuber storage protein in yam, has been reported to possess carbonic anhydrase, trypsin inhibitor, angiotensin-converting enzyme (ACE) inhibitor, free-radical scavenger, dehydroascorbate reductase and monodehydroascorbate reductase activities. Recent research has also found that dioscorin can enhance immune modulation via the toll-like receptor 4 (TLR-4) signal transduction pathway in RAW 264.7 cells, murine bone-marrow cells and human monocytes ex vivo.

Cloning of genes and enzymatic characterizations of novel dioscorin isoforms from Dioscorea japonica.

Dioscorin, the major tuber storage protein of yam, has been shown to possess carbonic anhydrase, trypsin inhibitor, dehydroascorbate reductase, and monodehydroascorbate reductase activities. In the present study, dioscorin from Dioscorea japonica was confirmed as a glycoprotein using the enhanced concanavalin A-peroxidase staining method, and the protein was shown to have both N- and O-glycans. Following the gene cloning, four full-length isoforms of dioscorin were expressed in Escherichia coli and purified by affinity purification and anion-exchange chromatography for structural and biochemical experiments.

Isolation and tyrosinase inhibitory effects of polyphenols from the leaves of persimmon, Diospyros kaki.

The main polyphenols were isolated from the leaves of six selected persimmon cultivars. Seven compounds were obtained by reverse-phase HPLC, and their structures were elucidated by multiple NMR measurements. These compounds are hyperoside, isoquercitrin, trifolin, astragalin, chrysontemin, quercetin-3-O-(2''-O-galloyl-β-D-glucopyranoside) (QOG), and kaempferol-3-O-(2''-O-galloyl-β-D-glucopyranoside) (KOG).

Isolation and Caenorhabditis elegans lifespan assay of flavonoids from onion.

The main flavonoids were isolated from three selected onion cultivars. Three phenolic compounds were obtained by reverse-phase HPLC, and their structures were elucidated by multiple NMR measurements. There were two known compounds, quercetin and quercetin 3'-O-β-D-glucopyranoside (Q3'G), and one novel compound, quercetin 3-O-β-D-glucopyranoside-(4→1)-β-d-glucopyranoside (Q3M), which was identified in onion for the first time.

One-week antihypertensive effect of Ile-Gln-Pro in spontaneously hypertensive rats.

The antihypertensive effect of an angiotensin I-converting enzyme (ACE) inhibitory peptide Ile-Gln-Pro (IQP), whose sequence was derived from Spirulina platensis , was investigated in spontaneously hypertensive rats (SHRs) for 1 week. The weighted systolic blood pressure (SBP) and diastolic blood pressure (DBP) of the peptide IQP-treated group were significantly lower than those of the negative control group from the third and fourth days, respectively. Accompanying the blood pressure reduction, a significant regulation of the expression of major components of the renin-angiotensin system (RAS) was found in the treatment group, including downregulation of the mRNA levels of renin, ACE, and the angiotensin II type 1 (AT1) receptor in the kidney, as well as serum angiotensinogen (Ang), ACE, and angiotensin II (Ang II) concentrations.

Isolation of an antihypertensive peptide from alcalase digest of Spirulina platensis.

An angiotensin I-converting enzyme (ACE) inhibitory peptide Ile-Gln-Pro with an IC(50) value of 5.77 +/- 0.09 microM was purified from the alcalase digests of Spirulina platensis by gel filtration chromatography and two steps of reverse-phase high-performance liquid chromatography (RP-HPLC).

Structural basis of abscisic acid signalling.

The phytohormone abscisic acid (ABA) mediates the adaptation of plants to environmental stresses such as drought and regulates developmental signals such as seed maturation. Within plants, the PYR/PYL/RCAR family of START proteins receives ABA to inhibit the phosphatase activity of the group-A protein phosphatases 2C (PP2Cs), which are major negative regulators in ABA signalling. Here we present the crystal structures of the ABA receptor PYL1 bound with (+)-ABA, and the complex formed by the further binding of (+)-ABA-bound PYL1 with the PP2C protein ABI1.

Intestinal uptake of amyloid beta protein through columnar epithelial cells in suckling mice.

The mechanism of transmission of amyloid protein, especially the dynamics in the intestine, is still largely unknown. In the present study, a fusion protein (Abeta-EGFP) that combined enhanced green fluorescent protein with amyloid-beta protein (Abeta) was orally administered to mice before and after weaning, and the uptake and kinetics of amyloid protein within the intestine were elucidated through histopathology. Abeta-EGFP was incorporated into the cytoplasm of columnar epithelial cells, rather than M cells, at 3 h after administration and thereafter.

Absolute side-chain structure at position 13 is required for the inhibitory activity of bromein.

Bromelain isoinhibitor (bromein), a cysteine proteinase inhibitor from pineapple stem, has a unique double-chain structure. The bromein precursor protein includes three homologous inhibitor domains, each containing an interchain peptide between the light and heavy chains. The interchain peptide in the single-chain precursor is immediately processed by bromelain, a target proteinase.

Arabidopsis DREB2A-interacting proteins function as RING E3 ligases and negatively regulate plant drought stress-responsive gene expression.

The DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN2A (DREB2A) transcription factor controls water deficit-inducible gene expression and requires posttranslational modification for its activation. The activation mechanism is not well understood; however, the stability of this protein in the nucleus was recently found to be important for its activation. Here, we report the isolation of Arabidopsis thaliana DREB2A-INTERACTING PROTEIN1 (DRIP1) and DRIP2, C3HC4 RING domain-containing proteins that interact with the DREB2A protein in the nucleus.

Proteinase inhibitor from ginkgo seeds is a member of the plant nonspecific lipid transfer protein gene family.

A 9-kD proteinase inhibitor was isolated from the seeds of ginkgo (Ginkgo biloba) and purified to homogeneity. This protein was revealed to partial-noncompetitively inhibit the aspartic acid proteinase pepsin and the cysteine proteinase papain (inhibition constant = 10(-5)-10(-4) m). The cDNA of the inhibitor was revealed to contain a 357-bp open reading frame encoding a 119-amino acid protein with a potential signal peptide (27 residues), indicating that this protein is synthesized as a preprotein and secreted outside the cells.

Crystallization and preliminary X-ray analysis of ginkbilobin-2 from Ginkgo biloba seeds: a novel antifungal protein with homology to the extracellular domain of plant cysteine-rich receptor-like kinases.

The antifungal protein ginkbilobin-2 (Gnk2) from Ginkgo biloba seeds does not show homology to other pathogenesis-related proteins, but does show homology to the extracellular domain of plant cysteine-rich receptor-like kinases. Native Gnk2 purified from ginkgo nuts and the selenomethionine derivative of recombinant Gnk2 (SeMet-rGnk2) were crystallized by the sitting-drop vapour-diffusion method using different precipitants. X-ray diffraction data were collected from Gnk2 at 2.