Abstract:The binding change mechanism for the ATP synthase has two central features. One is that the principle use of energy required for ATP synthesis is to promote the release of tightly bound ATP and the binding of Pi and ADP in a manner competent to form bound ATP. The second is that during net ATP formation multiple catalytic sites on the synthase participate in strongly cooperative sequence. Rotation of the γ subunit in F₁ is thought to deform the catalytic sites to give binding change. When the crystal structure of the F₁-ATPase was eventually solved, direct evidences for rotation of subunits during catalysis of F₁-ATPase were provided.

Abstract:DNA recognition of the transcription factors involves both general chemical rules and specific stereochemical rules. Recognition helice of transcription factor families binds to DNA, positing at a major groove. Pairing between the “residues line” of recognition helix and “base line” of base positions makes full fitting, usually involving 3 turns of α-helix and 3～5 base pairs.The binding geometry determined by interaction of the residues and bases in recognition area is indicated in the stereochemical chart, which shows recognition specificity. Stereochemical rules of DNA recognition by transcription factor family are summarized at bases of the chart.

Abstract:PNA is DNA analog in which the phosphate backbone has been replaced by (2-aminoethyl) glycine unit that is linked to the nucleotide bases via the glycine amino nitrogen and methylenecarbonyl linkers. PNA can bind to complementary oligonucleotides by Watson-Crick base paring with high thermal stability and exhibit wide biological effects including modulating the function of DNA sequence specific binding protein and modulating the transcription and translation in vivo or in vitro. Many applications have been explored for PNA as a new kind of molecular biological tools. Despite its DNA(RNA)binding properties, recent progress has shown that PNA has potential for the development of gene-targeting pharmaceuticals.

Abstract:There are two kinds of ubiquitous calpains, calpain Ⅰ and calpain Ⅱ, differing in their Ca²⁺ requirements for half maximum activities. Both calpains have a large subunit and a small subunit, with molecular weights of 80 and 30 ku respectively. Large subunit is composed of 4 domains. Small subunit is composed of 2 domains. Recently, several tissue specific calpains were discovered, adding to the complexity of calpain system. Calpastatin is an endogenous suppressor of calpain, which can bind to activated calpain specifically and made them inactive. There are 5 domains in calpastatin, domain L and 4 repeated domains numbered 1 to 4 which are responsible for their inhibity effects. Calpain activity is restrictively regulated in living cells. Membrane attachment reaction can low the Ca²⁺ requirement of calpain to be activated. The negatively charged phosphate groups on the polar head of membrane phospholipids are inportment for that activation. Autolysis also can low the Ca²⁺ requirement of calpain.

Abstract:SRY is the only gene currently known to be involved in the process of sex determination. A number of cloned genes probably participate in gonadal development: the MIS (also called AMH), the SOX9, the SF1, the DAX1, the WT1 and the DSS etc. Some gene model for mammalian sex determination such as Z-gene model and DSS-gene model etc. have recently been proposed. Those models provide a rational explanation not only for the cases of XX and XY sex reversal currently known to occur in humans and other mammals,but also for molecular mechanism of sex determination. Many questions of mammalian sex determination still remain unresolved. The identification and functional analysis of other genes in the mammalian sex determining cascade will determine the validity of this hypothesis.

Abstract:α-Amidation is a critical post-translational processing of many bioactive peptides in the nervous and endocrine systems. PAM, a bifunctional enzyme, with two catalytic domains PHM and PAL, catalyzes the sequential two-step conversion of glycine-extended peptides into COOH-terminal amidated peptides. Alternative splicing and tissue specific processing generate multiple forms of PAM. As a rate-limiting enzyme in biosynthetic pathway of peptides, levels of PAM are tissue specific and under the regulation of hormones and developmental cues.

Abstract:A brief introduction is given about the application of X-ray diffraction to the study of the dynamic process of the protein. Firstly, it gives some description about using traditional and Laue X-ray diffraction to study the dynamic process of a protein in a reaction taking place in a few minutes. Secondly, it is about reaction synchronization used to study the dynamic process of a protein in a few seconds. By choosing unmatchable substrate and unsuitable pH value, by choosing temperature and pH value jumping and by choosing metal ion and photochemical induction, enzyme reaction can be activated instantaneously.

Abstract:Tenascin-R (TN-R) is an important extracellular matrix glycoprotein. Tenascin-R is restricted to the central nervous system and mainly express in the early developing of oligodendrocytes. It also expreses in mature oligodendrocytes and some neural cell types (Such as spin cord. optic nerve, inter neuron of cerebellum and hippocampus).Tenascin-R has a modular structune with a cysteine-rich amino-terminal region followed by epidermal growth factor (EGF)-like repeats(EGF-L), fibronectin-typeⅢ homologous repeats(FNⅢ) and a fibrinogen like domain at the carboxy-terminal end.Tenascin-R has perplexing multifunctionality. Tenascin-R acts as a repellent molecule for neuron, promotes or inhibites the outgrowth of neurite,induces a polarity morphology of neurons and relates to myelinating processes. Tenascin-R has multiple receptors. F3/F11, XL1, Xprocan and MAG which have been isolated and characterized.

Abstract:The tryptophan fluorescence of a subunit of F_o moiety of mitochondrial F₁F_o can be quenched by the addition of Hypocrellin B. The determination of the Stern-Volmer plot at different temperatures is carried out. The result shows that the K_sv increased with the increase of temperature. The experimental result of the time-resolved fluorescence decay shows the decrease of lifetime of tryptophan fluorescence of F_o with the increase of concentration of HB. No shift in the absorbance spectra of F₁F_o were occurrent at varying concentration of HB. The results support the dynamic quenching. In addition, HB possess the necessary characters to be used as a quencher in the hydrophobic phase as follows: the low concentration of effective quenching; no effect on the activity of F₁F_o; the ratio of the partition coefficients between lipid-phase and water-phase as high as 16 560∶1. So HB can be used as a ideal fluorescence quencher for the study of conformational change of F_o moiety of membrane-bound F₁F_o complex.

Abstract:When conventional gel electrophoresis was used, it was found that DNA of NIH3T3 cells cultured for different time after UVB irradiation had no DNA ladders, but thymocytes of Kunming mouse processed by the same method had DNA ladder. The results of field inversion electrophoresis showed that after UVB irradiation, the DNA of NIH3T3 cleaved into high molecular weight fragments at first, and then into low molecular weight fragments, still without the appearance of DNA ladders. It is suggested that DNA cleavage of apoptotic cell is not always initiate at internucleosome.

Abstract:A newly identified gene MK, specifies for a heparin binding factor, is transiently expressed in the midgestation period of embryogenesis and in the early stages of embryonal carcinoma cell differentiation. In late embryos and the adult, MK gene is expressed only in the kidney. MK plays decisive roles not only in growth regulation but also in regulation of cell differentiation. The encoding sequence of mature peptide of MK was obtained by RT-PCR from human fetal kidney and was cloned into pBV221, after transfering into E.coli. A highly expressed vector of MK was constructed.

Abstract:In order to search the influence of Ⅰ-A^k_αβ gene transfer on the growth of the transferred mouse tumor cells in vivo, the α and β chain cDNA were synthesized, and inserted into a retroviral vector, to construct two Ⅰ-A^k expressing recombinant plasmids pLSXN-A^k_α and pLSXN-A^k_β. By means of a liposome-mediated gene transfer procedure, these two recombinant plasmids together were introduced into mouse lymphoma EL4 cells and mastocytoma P815 cells. The expression of the Ⅰ-A^k_αβ protein on the surface of the transconducted tumor cells was tested by FACS using Anti-A^k-FITC. Afterwards these tumor cells were injected subcutaneously into autologous mice C57BL/6(H-2^b) and DBA/2(H-2^d) respectively. It was observed that tumors developed at the beginning within the mouse bodies, and disappeared after a few weeks with injection. While the nontransferred tumor cells grew continuously. These results show that the tumor immune response can be stimulated by the tumor cells transferred with allogenic MHC class Ⅱ gene, in the absence of B7 co-stimulatory signals.

Abstract:The single cell gel electrophoresis (SCGE), also called comet assay, is a simple,rapid and sensitive biochemical technique for detecting DNA single strand breakage of the mammalian cells. In order to study the anti-cancer mechenism of CM4-component of the anti-bacterial peptides (CM4-ABP), the SCGE was used to observe the chromatin DNA breakage action of the K562 cancer cells comparing with the leucocytes of normal human under treating with CM4-ABP.Using the fluorescent microscopy, it is observed the chromatin DNA cleavage of the K562 cancer cells treated with CM4-ABP forms brighte fluorescent head and comet like tail; On the contrary, the normal human leucocytes treated and the K562 cancer cells untreated with CM4-ABP show intact, round nuclei, no comet tails. The analysis of comet assay showed that the averge ratio of the K562 nuclei cleavage is 73.62% (P＜0.001). The results showed that the CM4-ABP can cause the chromatin DNA of the K562 cells damaged, but it has no normal human leucocytes.

Abstract:The expression vector was successfully constructed with the cloning of erythropoietin genomic DNA into the downstream of CMV promoter,and expressed in transfected transiently BHK21 cells. Then it was introduced into the skeletal muscle of mice by the mediation of lipofectin. As a result, the EPO expressed in serum of mice reached highly to 1 340 ng/L in one month.

Abstract:An enzyme-amplified lanthanide luminescence assay of alkaline phosphatase was described. Optimization of important factors in this assay was studied. Improvement of stability of signal/noise ratio by using methyl silicon oil (I) was first reported. The sensitivity and the accuracy of the assay were 4 U/L and ＜10% respectively. The concentration range corresponding to 10% precision was 2.00～3.02×10² U/L. The activity of alkaline phosphatase in serum was also detected and the recovery was 93%～95%. 5-fluorosalicyl phosphate was synthesized and used as substrate.

Abstract:Reverse dot blot (RDB) technique was used. Seven sequence-specific oligonucleotide (SSO) probes directed against 7 types of HPV (HPV6B, 11, 16, 18, 31, 33 and 35) respectively are synthesized and fixed sequentially on a stripe of nylon membrane to form 7 spots. After hybridization with the PCR product of the sample DNA sequence, any one of the 7 types of HPV DNA were distinguished on only one stripe of nylon membrane. 38 cases of cervical carcinoma samples were detected by this PCR-RDB technique. The results show: 29 cases (76.3%) of HPV16 positive; 8 cases (21.1%) of HPV18 positive; 2 cases (7.2%)of double infection; and 3 cases (8.0%) of HPV negative.The method is rapid and simple, with high specificity, no false positive results in general, and as the PCR technique is very sensitive, the method is not liable to show false negative results.

Abstract:Soluble-insoluble forms of monoclonal antibody, which can be rendered either soluble or insoluble by simple adjusting the temperature or pH, have been prepared by covalently coupling antibody to poly(N-isopropylacrylamide) or to copolymer of methacrylic acid and acrylamide. These carriers were used in enzyme immunoassay instead of polystyrenes. Some advantages of both heterogeneous and homogeneous immunoassays were achieved. HBsAg was assayed to examine the sensitivity of these methods. A significant difference from the negative serum was obtained for 0.5 μg/L. 43 serum samples were assayed, and there was a good correlation between these methods and ELISA.

Abstract:Genomic DNA of human placenta chorionic cells and leukocytes were hybridized with Emx probe which is a cDNA fragment including homeobox from quail. The results showed Emx gene dose in human placenta chorioic cells was six-fold more than in leukocytes and indicated Emx gene amplified in human placenta chorioic cells.

Abstract:The detection of DNA cross-link induced by diaminedichloroplatinum (CDDP) in HeLa line irradiated with ⁶⁰Co γ-ray was carried out using ethidium bromide fluorescence assay. The results showed that after single doses (1～6 Gy)of irradiation, the fraction of control growth of HeLa cells declined and the DNA content per unit cell of HeLa line increased, and the formation of interstrand DNA cross-link induced by CDDP reduced apparently with increments of dose of irradiation. The findings suggest that tumor cells surviving γ-irradiation may become resistant to chemotherapy with alkylating agents such as CDDP.

Abstract:A recombinant expression plasmid was constructed by inserting the cDNA fragment encoding human sperm membrane protein (HSDⅡ) into pGEX vector. High level expression of the fusion protein was observed in the DH5α bacteria transformed with the recombinant pGEX vector in the absence of IPTG as well as after IPTG induction according to PAGE detection. A simple and valid method was recommended to minimize the basal level expression without IPTG induction: a plasmid pREP4 carrying LacⅠ gene was cotransformed with the recombinant expression plasmid into DH5α, and the basal level expression of the fusion protien was inhibited significantly while the presense of pREP4 did not affect overall expression following induction with IPTG.