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91.
The N-terminal -amino groups of 1-bungarotoxin (1-Bgt) fromBungarus multicinctus venom were modified with trinitrobenzene sulfonic acid and the modified derivative was separated by high performance liquid chromatography. The trinitrophenylated (TNP) derivative contained two TNP groups at the -amino groups of A chain and B chain and showed a marked decrease in enzymatic activity. Methionine residues at positions 6 and 8 of the A chain were oxidized with chloramine T or cleaved with cyanogen bromide to remove the N-terminal octapeptide. Oxidation of methionine residues and removal of the N-terminal octapeptide caused a precipitous decrease in enzymatic activity, whereas antigenicity remained unchanged. The presence of dihexanoyllecithin influenced the interaction between 1-Bgt and 8-antilinonaphthalene sulfonate (ANS) and revealed that 1-Bgt consists of two types of ANS-binding sites, one at the substrate binding site of the A chain and the other might be at the B chain. The modified derivatives still retained their affinity for Ca2+ and ANS, indicating that the N-terminal region is not involved in Ca2+ and substrate binding. A fluorescence study revealed that the -amino group of the A chain was in the vicinity of substrate binding site and that the TNP -amino groups were in proximity to Trp-19 of the A chain. In addition, the study showed that the N-terminal region is important for stabilizing the architectural environment of Trp-19. The results, together with the proposal that Trp-19 of the A chain is involved in substrate binding, suggest that the N-terminal region of the A chain plays a crucial role in maintaining a functional active site for 1-Bgt. 相似文献
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93.
水稻原生质体产生细胞团的冰冻保存和冻后再生植株形成 总被引:4,自引:0,他引:4
水稻(Oryza sativa L.)原生质体产生的细胞团加上10-20%的二甲亚枫(DMSO)和10-20%的蔗糖,置于液氮中保存。冻后细胞生存率达到对照的40-50%。存活的细胞在附加2×10~(-5)mol/l 2,4-D 的Linsmier-Skoog(Ls)固体培养基上再生长,然后将形成的愈伤组织块转到附加10~(-6)mol/l NAA,4×10~(-6)mol/l 激动素和10~(-6)mol/l 2 IP 及8%的蔗糖的 LS培养基上分化出芽并形成植株。 相似文献
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96.
Experiments were conducted to develop a simple and reliable technique to produce chimeric rabbits from morula stage embryos. In Experiments 1 and 2, an in-vitro test of viability was initially performed by culturing embryos to the blastocyst stage. Ninety-three percent of the “chimeric” embryos developed to the blastocyst stage compared to 94% for controls when embryos were manipulated soon after collection (Exp. 1). Eighty-one percent chimeric embryos and 78% control embryos developed to blastocyst stage when embryos were held at room temperature for 4 hr (Exp. 2). In Experiment 3, enough morula-stage embryos were available from true breeding Dutch-belted and albino rabbits to form potentially 67 diverse “color” pairs. These micromanipulated pairs of morulae were successfully combined to produce 64 chimeric embryos (96%, 64/67). They were transferred to the uteri of seven recipient does and three became pregnant producing 13 young. Four of the young exhibited substantial overt chimerism (31%) and one more was a possible chimera. 相似文献
97.
Shiaw-Der Yang Jen-Shin Song Yao-Tsung Hsieh Hui-Wen Liu Wen-Hsiung Chan 《Journal of Protein Chemistry》1992,11(5):539-546
The ATP·Mg-dependent protein phosphatase activating factor (Fa) has been identified and purified to near homogeneity from brain. In this report, as evidenced on SDS-polyacrylamide gel electrophoresis followed by autoradiography, factorFa has further been identified as a cAMP and Ca2+-independent brain kinase that could phosphorylate synapsin I, a neuronal protein that coats synaptic vesicles, binds to cytoskeleton, and is believed to be involved in the modulation of neurotransmission. Kinetic study further indicated that factorFa could phosphorylate synapsin I with a lowK
m
value of about 2 µM and with a molar ratio of 1 mol of phosphate per mole of protein. Peptide mapping analysis revealed that factorFa specifically phosphorylated the tail region of synapsin I but on a unique site distinct from those phosphorylated by Ca2+/calmodulin-dependent protein kinase II and cAMP-dependent protein kinase, the two well-established synapsin I kinases. Functional study further revealed that factorFa could phosphorylate this unique specific site on the tail region of synapsin I and thereby inhibit cross-linking of synapsin I with microtubules. The results further suggest the possible involvement of factorFa as a synapsin I kinase in the regulation of axonal transport process of synaptic vesicles via the promotion of vesicles motility during neurotransmission. 相似文献
98.
Protein phosphorylation was studied in Xanthomonas campestris pv. oryzae in vivo and in vitro. In vitro labelling showed that the protein kinases in this bacterium used both ATP and GTP as nucleotide substrates at nearly the same efficiency. At least 6 proteins were phosphorylated in vitro, including abundant species of p81, p44, and p32 with M
r of 81000, 44000, and 32000, respectively. Three types of phosphate-protein linkage were found in this bacterium: O-phosphate, N-phosphate and probably acyl phosphate. The p81 and p32 were phosphorylated at histidine. The p44 had mainly phosphoserine and a small part of phosphohistidine. The phosphorylation profile was variable depending on the growth conditions. Furthermore, by a virulent phage Xp10 infection the quantity of phosphorylation increased: for phosphohistinine more than 10-fold, and for phosphoserine about 3-fold. Thus, in this bacterium phosphorylation may be linked with a physiological regulation system and with Xp10 phage development. 相似文献
99.
Molecular cloning of the cDNA for the catalytic subunit of human DNA polymerase delta. 总被引:3,自引:0,他引:3
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C L Yang L S Chang P Zhang H Hao L Zhu N L Toomey M Y Lee 《Nucleic acids research》1992,20(4):735-745
The cDNA of human DNA polymerase delta was cloned. The cDNA had a length of 3.5 kb and encoded a protein of 1107 amino acid residues with a calculated molecular mass of 124 kDa. Northern blot analysis showed that the cDNA hybridized to a mRNA of 3.4 kb. Monoclonal and polyclonal antibodies to the C-terminal 20 residues specifically immunoblotted the human pol delta catalytic polypeptide. A multiple sequence alignment was constructed. This showed that human pol delta is closely related to yeast pol delta and the herpes virus DNA polymerases. The levels of pol delta message were found to be induced concomitantly with DNA pol delta activity and DNA synthesis in serum restimulated proliferating IMR90 cultured cells. The human pol delta gene was localized to chromosome 19 by Southern blotting of EcoRI digested DNA from a panel of rodent/human cell hybrids. 相似文献
100.