Background: Human Heparansulphate interacting protein (hHip) has been shown to participate in biological processes of cells. Several studies indicated that hHip transcript is up regulated in several of cancer tissues including those of thyroid, colon, breast and prostate. Antibody against hHIP is necessary for methods to evaluate protein level of HIP in cancer tissues. Objectives:The aims of study was to induce anti hHIP antibody in rabbit and purify and conserve purified anti hHIP antibody. Subjects and method: The study included 9 adult and healthy rabbits with the weight 2 - 2.5kg. Immunization hHIP peptide-KLH in rabbit. Purify anti hHIP antibody using affinity chromatography. Results: The results shown synthesize hHIP peptide and conjugate it with carrier protein. Sensitive rabbit better meet with hHIP-KLH antibody. The Ig concentration obtained in sensitive rabbit was rather high and equal. Immunization hHIP-KLH successfully in rabbit. Obtainment valuable amount of anti hHIP antibody. Conclusion: Successfully induce and purify anti hHIP antibody from rabbit. Establish a standard protocol for polyclonal antibody against small peptide in rabbit.\r\n', u'\r\n', u'
Carrier Proteins/ administration & ; dosage ; chemistry ; Rabbits

PICK1 (protein interacting with C kinase 1) contains a PDZ (PSD-95/Dlg/ZO1) domain and a BAR (Bin/amphiphysin/Rvs) domain. Via the PDZ domain, PICK1 interacts directly with more than 40 proteins. Among these interacting proteins, some are important for physiological and pathophysiological processes of central nervous system. In this review, recent findings about how PICK1 is associated with central nervous system diseases are summarize.
Animals ; Carrier Proteins ; chemistry ; metabolism ; physiology ; Epilepsy ; metabolism ; Humans ; Nuclear Proteins ; chemistry ; metabolism ; physiology ; Schizophrenia ; metabolism ; Stroke ; metabolism

This article reported the clinical features of one child with infantile hypophosphatasia (HPP) and his pedigree information. The proband was a 5-month-old boy with multiple skeletal dysplasia (koilosternia, bending deformity of both radii, and knock-knee deformity of both knees), feeding difficulty, reduction in body weight, developmental delay, recurrent pneumonia and respiratory failure, and a significant reduction in blood alkaline phosphatase. Among his parents, sister, uncle, and aunt (other family members did not cooperate with us in the examination), his parents and aunt had a slight reduction in alkaline phosphatase and his aunt had scoliosis; there were no other clinical phenotypes or abnormal laboratory testing results. His ALPL gene mutation came from c.228delG mutation in his mother and c.407G>A compound heterozygous mutation in his father. His aunt carried c.228delG mutation. The c.407G>A mutation had been reported as the pathogenic mutation of HPP, and c.228delG mutation was a novel pathogenic mutation. Hypophosphatasia is caused by ALPL gene mutation, and ALPL gene detection is an effective diagnostic method. This study expands the mutation spectrum of ALPL gene and provides a theoretical basis for genetic diagnosis of this disease.
Alkaline Phosphatase ; genetics ; Carrier Proteins ; chemistry ; Female ; Heterozygote ; Humans ; Hypophosphatasia ; etiology ; genetics ; Infant ; Male ; Mutation ; Pedigree

Carrier Proteins ; genetics ; Cloning, Molecular ; DNA, Complementary ; Humans ; Liver ; chemistry ; Membrane Glycoproteins ; genetics

OBJECTIVESTo detect the cholesterol ester transfer protein (CETP) levels in semen of infertile patients and evaluate the correlation between CETP in semial plasma and infertility.

METHODSOne hundred and sixty-three infertile patients and fifteen fertile males were selected randomly. The routine examination of ejaculates was fulfilled by computer aided semen analysis (CASA). The CETP levels in all seminal plasma samples and fifty-five serum samples were detected by ELISA method.

RESULTSThe CETP levels in infertile patients and fertile males were (2.21 +/- 1.23) microgram/L and (1.40 +/- 0.45) microgram/L, respectively. There were no significant differences between the two groups(P > 0.05). And there were no significant differences of CETP levels in seminal plasma among groups of azoospermia(n = 29), oligoasthenozoospermia (n = 58), oligospermia(n = 15), asthenozoospermia(n = 44) and normozoospermia(n = 17) in the infertile patients(P > 0.05). The CETP in seminal plasma and serum were detected in 55 infertile patients, and there was no correlation between CETP levels in seminal plasma and serum using Spearman analysis(r = 0.009, P > 0.05). The mean CETP level in seminal plasma was almost 1/1,000 of that in serum.

CONCLUSIONSThe CETP level in seminal plasma is extremely low and has no relation with the changes of sperm density or motility. It may ensure the integrity of sperm membrane before the sperm enters into female genital tract.

Adult ; Carrier Proteins ; analysis ; blood ; Cholesterol Ester Transfer Proteins ; Glycoproteins ; Humans ; Infertility, Male ; metabolism ; Male ; Middle Aged ; Semen ; chemistry

To prepare anti-mouse uteroglobin binding protein (mUGBP) polyclonal antibody, two polypeptides were synthesized based on the bioinformatics analysis of mUGBP, and New Zealand white rabbits were immunized separately with each peptide coupled with keyhole limpet hemocyanin (KLH). The data indicate that a 13-amino acid polypeptide (positions 221st-233rd) was able to generate anti-peptide antibodies. The titer of the antisera detected with ELISA was 1:10(8). The antisera were then purified with immuno-affinity chromatography to obtain antibodies. Western blot analysis of mUGBP expressed as a fusion protein with a green fluorescent protein (GFP) was performed on the cell lysates of COS-1 cells with the purified antisera, suggesting that the antisera specifically recognized UGBP. By immunohistochemistry and indirect immunofluorescence analysis, we examined the expression of UGBP in the lung tissues from a patient undergoing surgical lung resection for a tumor and from normal mouse lung tissue, and found for the first time that UGBP protein was widely expressed in both mouse and human lung tissue with the most abundant expression in bronchial epithelial cells. These results suggest that the antigen epitopes of mUGBP are well predicted by using bioinformatics analysis. We have obtained anti-mUGBP polyclonal antibody, which will be useful for further investigation.
Animals ; Antibodies ; chemistry ; COS Cells ; Carrier Proteins ; chemistry ; Cercopithecus aethiops ; Computational Biology ; Enzyme-Linked Immunosorbent Assay ; Hemocyanins ; Humans ; Immune Sera ; Immunohistochemistry ; Mice ; Rabbits ; Recombinant Proteins ; chemistry ; Uteroglobin

This article is to summarize the molecular and functional analysis of the gene "suppression of tumorigenicity 13" (ST13). ST13 is in fact the gene encoding Hsp70 interacting protein (Hip), a co-factor (co-chaperone) of the 70-kDa heat shock proteins (Hsc/Hsp70). By collaborating with other positive co-factors such as Hsp40 and the Hsp70-Hsp90 organizing protein (Hop), or competing with negative co-factors such as Bcl2-associated athanogen 1 (Bag1), Hip facilitates may facilitate the chaperone function of Hsc/Hsp70 in protein folding and repair, and in controlling the activity of regulatory proteins such as steroid receptors and regulators of proliferation or apoptosis. Although the nomenclature of ST13 implies a role in the suppression of tumorigenicity (ST), to date available experimental data are not sufficient to support its role in cancer development, except for the possible down-regulation of ST13 in gastric and colorectal cancers. Further investigation of this gene at the physiological level would benefit our understanding of diseases such as endocrinological disorders, cancer, and neurodegeneration commonly associated with protein misfolding.
Adenosine Triphosphate ; metabolism ; Animals ; Carrier Proteins ; chemistry ; genetics ; physiology ; Cloning, Molecular ; HSP70 Heat-Shock Proteins ; metabolism ; Humans ; Protein Folding ; Tumor Suppressor Proteins ; chemistry ; genetics ; physiology

OBJECTIVETo investigate the role of 83 site in interaction of GluR2 C-terminal and PICK1 PDZ domain.

METHODSDocking structure of PICK1 PDZ domain with GluR2 C terminal PDZ binding motif was built with computer software. After K83 site was substituted by other amino acid, the structure and binding energy were recalculated; meanwhile, site specific mutants were constructed using wild type full length cDNA as template. Mutants were co-transfected with GluR2 into HEK293T cells. After staining, the distribution of PICK1 and GluR2 were observed under confocal microscope.

RESULTSWild type PICK1 and GluR2 formed many co-clusters in HEK293T cells as reported by other research groups; but different K83 mutant had different distribution in HEK293T cells.

CONCLUSIONThe K83 site in PDZ domain of PICK1 is important for the interaction between PICK1 and GluR2. Altering lysine will probably change the hydrophobic interactions, the hydrogen bonds or the electrostatic interactions formed between PICK1 PDZ domain and GluR2 C terminal; accordingly, that will change the binding capacity between PICK1 and GluR2 in varying degrees.

Binding Sites ; Carrier Proteins ; chemistry ; metabolism ; Computer Simulation ; HEK293 Cells ; Humans ; Nuclear Proteins ; chemistry ; metabolism ; PDZ Domains ; Protein Binding ; Receptors, AMPA ; metabolism

Faldaprevir analogue molecule (FAM) has been reported to effectively inhibit the catalytic activity of HCV NS3/4A protease, making it a potential lead compound against HCV. A series of HCV NS3/4A protease crystal structures were analyzed by bioinformatics methods, and the FAM-HCV NS3/4A protease crystal structure was chosen for this study. A 20.4 ns molecular dynamics simulation of the complex consists of HCV NS3/4A protease and FAM was conducted. The key amino acid residues for interaction and the binding driving force for the molecular recognition between the protease and FAM were identified from the hydrogen bonds and binding free energy analyses. With the driving force of hydrogen bonds and van der Waals, FAM specifically bind to the active pocket of HCV NS3/4A protease, including V130-S137, F152-D166, D77-D79 and V55, which agreed with the experimental data. The effect of R155K, D168E/V and V170T site-directed mutagenesis on FAM molecular recognition was analyzed for their effect on drug resistance, which provided the possible molecular explanation of FAM resistance. Finally, the system conformational change was explored by using free energy landscape and conformational cluster. The result showed four kinds of dominant conformation, which provides theoretical basis for subsequent design of Faldaprevir analogue inhibitors based on the structure of HCV NS3/4A protease.
Antiviral Agents ; chemistry ; Carrier Proteins ; chemistry ; Drug Resistance, Viral ; Endopeptidases ; Hepacivirus ; Molecular Dynamics Simulation ; Mutagenesis, Site-Directed ; Oligopeptides ; chemistry ; Protease Inhibitors ; chemistry ; Serine Proteases ; Thiazoles ; chemistry ; Viral Nonstructural Proteins ; chemistry

Pulldown assay is an in vitro method for studies of protein-protein interactions, in which tagged proteins are usually expressed as the bait to enrich other proteins that could bind to them. In this technology, the GST tag is broadest used for its modest size and hydrophilic property. In most cases, the GST tag could increase the hydrophility of the fusion protein and help to avoid the formation of inclusion bodies. However, in the other few cases, the target protein may be strongly hydrophobic or have complicated structures that were hard to fold and assemble in correct conformations without champerons, and even the existence of GST tag could not make them soluble. These proteins were always expressed as inclusion bodies and had no functions. LMO2 was a small molecular weight and insoluble protein, in this study, GST system and MBP system were used to express GST-LMO2 and MBP-LMO2 fusion proteins, respectively. We found that GST-LMO2 fusion protein was expressed as inclusion bodies whereas MBP-LMO2 fusion protein was expressed in soluble form. Moreover, the production rate of MBP-LMO2 was also much higher than GST-LMO2. Then MBP-LMO2 fusion proteins and renatured GST-LMO2 fusion proteins were used as bait in pulldown assay to study the interaction between LMO2 and endogenous GATA1 in K562 cells. Western blot analyses showed that both of these proteins could bind to endogenous GATA1 in K562 cells, but recovered GATA1 protein by MBP-LMO2 fusion protein was much more than GST-LMO2 fusion protein. These results suggest that using of MBP system is a helpful attempt in the case of studying small molecular weight, strong hydrophobic proteins.
Adaptor Proteins, Signal Transducing ; Carrier Proteins ; chemistry ; Chemical Precipitation ; DNA-Binding Proteins ; chemistry ; GATA1 Transcription Factor ; chemistry ; Genetic Vectors ; Glutathione Transferase ; chemistry ; Humans ; K562 Cells ; LIM Domain Proteins ; Maltose-Binding Proteins ; Metalloproteins ; chemistry ; Protein Binding ; Protein Interaction Domains and Motifs ; Protein Renaturation ; Proto-Oncogene Proteins ; chemistry ; Recombinant Fusion Proteins ; genetics ; metabolism