The effect of antibacterial peptide CM4 of Bombyx mori against E. coll K12 was investigated using scanning electron microscopy(SEM) and transmission electron microscopy (TEM). The ultrastructural changes of E. coli K1...The effect of antibacterial peptide CM4 of Bombyx mori against E. coll K12 was investigated using scanning electron microscopy(SEM) and transmission electron microscopy (TEM). The ultrastructural changes of E. coli K12 were observed by the challenge of the purified antibacterial peptide CM4. The results showed that the antibacterial peptide caused a series of pathological changes on E. coli. SEM and TEM revealed aggregates of bacteria and SEM revealed wrin-kled bacterial surfaces in the early stage. Thereafter, plasmolysis was observed with irregular holes appearing in the two ends of bacteria and the cytoplasmic contents of the cells leaking out. Finally, bacteria became empty vesicles and disintegrated into small fragments subsequently. Comparatively, the bacterial membrane was normal and the bacterial structure remained intact in the control group.展开更多
In order to establish a possible evolutionary correlation between α lactalbumin and lysozyme,site\|directed mutagenesis on α\|lactalbumin cDNA was performed to change α\|lactalbumin into lysozyme.The gene of α\|la...In order to establish a possible evolutionary correlation between α lactalbumin and lysozyme,site\|directed mutagenesis on α\|lactalbumin cDNA was performed to change α\|lactalbumin into lysozyme.The gene of α\|lactalbumin was cloned into phagemid pSK +vector and the single strand template of which was prepared.Thereafter,the mutagenic oligonucleotide primers were designed and used to synthesize the mutated double strand DNA.By means of this site\|directed mutagenesis,the amimo acid residues at position His 32 and Thr 33 of bovine α\|lactalbumin cDNA were substituted with Leu and Glu,respectively.Thus,the catalytic site of lysozyme was obtained in α\|lactalbumin.Furthermore,Tyr 103 was substituted by Ala and the lysozyme substrate binding conformation was formed in α\|lactalbumin as well.The structural as well as functional relationship between α\|lactalbumin and lysozyme indicated that there existed possible evolutionary correlation between the two proteins.展开更多
Glutamate synthase (EC 1 4 1 14) was purified to homogeneity from a cell\|free extract of Streptomyces lincolnensis by precipitation with streptomycin sulfate and ammonium sulfate, and column chromatography on DEAE\| ...Glutamate synthase (EC 1 4 1 14) was purified to homogeneity from a cell\|free extract of Streptomyces lincolnensis by precipitation with streptomycin sulfate and ammonium sulfate, and column chromatography on DEAE\| cellulose, Sepharose 6B, DEAE\|sephadex A\|50, hydroxyapatite and Sephadex G\|150. The enzyme activity is stabilized by addition of α ketoglutarate, PMSF,EDTA, β mercaptoethanol and glycerol. The native enzyme has a molecular weight of 138 000 and is composed of two nonidentical subunits with molecular weights of 81 000 and 57 000. Spectroscopic examination of the enzyme gave absorption maximum at 280 and none at 380 and 440 nm, indicating the absence of iron and flavin. The enzyme shows optimum activity at pH 7.2 and 30℃. Km values for α ketoglutarate, L\|glutamine and NADH were 417, 435, and 52.1 μmol/L, respectively. When NADPH was substituted for NADH as reductant, there was approximately 13% of the control activity. The activity of this glutamate synthase is inhibited by its products (i.e. glutamate and NAD), several metal ions, amino acids and tricarboxylic acid cycle intermediates.展开更多
文摘The effect of antibacterial peptide CM4 of Bombyx mori against E. coll K12 was investigated using scanning electron microscopy(SEM) and transmission electron microscopy (TEM). The ultrastructural changes of E. coli K12 were observed by the challenge of the purified antibacterial peptide CM4. The results showed that the antibacterial peptide caused a series of pathological changes on E. coli. SEM and TEM revealed aggregates of bacteria and SEM revealed wrin-kled bacterial surfaces in the early stage. Thereafter, plasmolysis was observed with irregular holes appearing in the two ends of bacteria and the cytoplasmic contents of the cells leaking out. Finally, bacteria became empty vesicles and disintegrated into small fragments subsequently. Comparatively, the bacterial membrane was normal and the bacterial structure remained intact in the control group.
文摘In order to establish a possible evolutionary correlation between α lactalbumin and lysozyme,site\|directed mutagenesis on α\|lactalbumin cDNA was performed to change α\|lactalbumin into lysozyme.The gene of α\|lactalbumin was cloned into phagemid pSK +vector and the single strand template of which was prepared.Thereafter,the mutagenic oligonucleotide primers were designed and used to synthesize the mutated double strand DNA.By means of this site\|directed mutagenesis,the amimo acid residues at position His 32 and Thr 33 of bovine α\|lactalbumin cDNA were substituted with Leu and Glu,respectively.Thus,the catalytic site of lysozyme was obtained in α\|lactalbumin.Furthermore,Tyr 103 was substituted by Ala and the lysozyme substrate binding conformation was formed in α\|lactalbumin as well.The structural as well as functional relationship between α\|lactalbumin and lysozyme indicated that there existed possible evolutionary correlation between the two proteins.
文摘Glutamate synthase (EC 1 4 1 14) was purified to homogeneity from a cell\|free extract of Streptomyces lincolnensis by precipitation with streptomycin sulfate and ammonium sulfate, and column chromatography on DEAE\| cellulose, Sepharose 6B, DEAE\|sephadex A\|50, hydroxyapatite and Sephadex G\|150. The enzyme activity is stabilized by addition of α ketoglutarate, PMSF,EDTA, β mercaptoethanol and glycerol. The native enzyme has a molecular weight of 138 000 and is composed of two nonidentical subunits with molecular weights of 81 000 and 57 000. Spectroscopic examination of the enzyme gave absorption maximum at 280 and none at 380 and 440 nm, indicating the absence of iron and flavin. The enzyme shows optimum activity at pH 7.2 and 30℃. Km values for α ketoglutarate, L\|glutamine and NADH were 417, 435, and 52.1 μmol/L, respectively. When NADPH was substituted for NADH as reductant, there was approximately 13% of the control activity. The activity of this glutamate synthase is inhibited by its products (i.e. glutamate and NAD), several metal ions, amino acids and tricarboxylic acid cycle intermediates.
