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2.9 A resolution structure of an anti-dinitrophenyl-spin-label monoclonal antibody Fab fragment with bound hapten 总被引:5,自引:0,他引:5
The crystal structure of the Fab fragment of the murine monoclonal anti-dinitrophenyl-spin-label antibody AN02 complexed with its hapten has been solved at 2.9 A resolution using a novel molecular replacement method. Prior to translation searches, a large number of the most likely rotation function solutions were subjected to a rigid body refinement against the linear correlation coefficient between intensities of observed and calculated structure factors. First, the overall orientation of the search model and then the orientations and positions of the four Fab domains (VH, VL, CH1 and CL) were refined. This procedure clearly identified the correct orientation of the search model. The refined search model was then subjected to translation searches which unambiguously determined the enantiomer and position in the unit cell of the crystal. The successful search model was refined 2.5 A crystal structure of the Fab fragment of HyHel-5 from which non-matching residues in the variable domains had been removed. HyHel-5 is a murine monoclonal antibody whose heavy and light chains are of the same subclass (gamma 1, kappa, respectively) as AN02. After molecular replacement the structure of the AN02 Fab has been refined using simulated annealing in combination with model building and conjugate gradient refinement to a current crystallographic R-factor of 19.5% for 12,129 unique reflections between 8.0 and 2.9 A. The root-mean-square (r.m.s.) deviation from ideal bond lengths is 0.014 A, and the r.m.s. deviation from ideal bond angles is 3.1 degrees. The electron density reveals the hapten sitting in a pocket formed by the loops of the complementarity determining region. The dinitrophenyl ring of the hapten is sandwiched between the indole rings of Trp96 of the heavy-chain and Trp91 of the light-chain. The positioning of the hapten and general features of the combining site are in good agreement with the results of earlier nuclear magnetic resonance experiments. 相似文献
4.
Population genetics and phylogenetics of DNA sequence variation at multiple loci within the Drosophila melanogaster species complex 总被引:13,自引:1,他引:13
Two regions of the genome, a 1-kbp portion of the zeste locus and a 1.1-
kbp portion of the yolk protein 2 locus, were sequenced in six individuals
from each of four species: Drosophila melanogaster, D. simulans, D.
mauritiana, and D. sechellia. The species and strains were the same as
those of a previous study of a 1.9-kbp region of the period locus. No
evidence was found for recent balancing or directional selection or for the
accumulation of selected differences between species. Yolk protein 2 has a
high level of amino acid replacement variation and a low level of
synonymous variation, while zeste has the opposite pattern. This contrast
is consistent with information on gene function and patterns of codon bias.
Polymorphism levels are consistent with a ranking of effective population
sizes, from low to high, in the following order: D. sechellia, D.
melanogaster, D.mauritiana, and D. simulans. The apparent species
relationships are very similar to those suggested by the period locus
study. In particular, D. simulans appears to be a large population that is
still segregating variation that arose before the separation of D.
mauritiana and D. sechellia. It is estimated that the separation of
ancestral D. melanogaster from the other species occurred 2.5-3.4 Mya. The
separations of D. sechellia and D. mauritiana from ancestral D. simulans
appear to have occurred 0.58- 0.86 Mya, with D. mauritiana having diverged
from ancestral D. simulans 0.1 Myr more recently than D. sechellia.
相似文献
5.
Background
Elucidation of the communal behavior of microbes in mixed species biofilms may have a major impact on understanding infectious diseases and for the therapeutics. Although, the structure and the properties of monospecies biofilms and their role in disease have been extensively studied during the last decade, the interactions within mixed biofilms consisting of bacteria and fungi such as Candida spp. have not been illustrated in depth. Hence, the aim of this study was to evaluate the interspecies interactions of Pseudomonas aeruginosa and six different species of Candida comprising C. albicans, C. glabrata, C. krusei, C. tropicalis, C. parapsilosis, and C. dubliniensis in dual species biofilm development. 相似文献6.
Characterization of compost produced from separated pig manure and a variety of bulking agents at low initial C/N ratios 总被引:1,自引:0,他引:1
Nolan T Troy SM Healy MG Kwapinski W Leahy JJ Lawlor PG 《Bioresource technology》2011,102(14):7131-7138
The aim of this study was to investigate the composting of separated pig manure solids with or without a variety of bulking agents at a low initial C/N ratio (12.5-23.3). Compost stability was investigated using an oxygen uptake rate (OUR) test and compost maturity was investigated using a germination index test. All treatments showed typical patterns of compost temperature. Temperatures above 60 °C were achieved by Day 2, followed by a thermophilic phase (50-60 °C), which lasted for 1 to 2 weeks followed by a cooling phase. The stability of one of treatments which did not contain any bulking agent - OUR of 25 mmol O2 kg−1 OM hour−1 - was negatively affected by its initial high water content (69%). The addition of a bulking agent and initial water content below 60% were necessary to compost the separated solid fraction of pig manure at a low initial C/N ratio. 相似文献
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8.
Background
There have been many algorithms and software programs implemented for the inference of multiple sequence alignments of protein and DNA sequences. The "true" alignment is usually unknown due to the incomplete knowledge of the evolutionary history of the sequences, making it difficult to gauge the relative accuracy of the programs. 相似文献9.
10.
Leahy M Vaughan P Fanning L Fanning S Sheehan D 《Enzyme and microbial technology》2001,28(7-8):682-688
A recombinant Rhizobium meliloti beta-galactosidase was purified to homogeneity from an Escherichia coli expression system. The gene for the enzyme was cloned into a pKK223-3 plasmid which was then used to transform E. coli JM109 cells. The enzyme was purified 35-fold with a yield of 34% by a combination of DEAE-cellulose (pH 8.0) and two sequential Mono Q steps (at pH 8.0 and 6.0, respectively). The purified enzyme had an apparent molecular mass of 174 kDa and a subunit molecular weight of 88 kDa, indicating that it is a dimer. It was active with both synthetic substrates p-nitrophenyl beta-D-galactopyranoside (PNPG) and o-nitrophenyl beta-D-galactopyranoside (ONPG) with K(m)(PNPG) and K(m)(ONPG) of 1 mM at 25 degrees C. The k(cat)/K(m) ratios for both substrates were approximately 70 mM(-1) sec(-1), indicating no clear preference for either PNPG or ONPG, unlike E. coli beta-galactosidase. After non-denaturing electrophoresis, active beta-galactosidase bands were identified using 5-bromo-4-chloro-3-indolyl beta-D-galactopyranoside (X-gal) or 6-bromo-2-naphthyl beta-D-galactopyranoside (BNG) and diazo blue B. 相似文献