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1.
We have investigated the time course of the degradation of a supported dipalmitoylphosphatidylcholine bilayer by phospholipase A2 in aqueous buffer with an atomic force microscope. Contact mode imaging allows visualization of enzyme activity on the substrate with a lateral resolution of less than 10 nm. Detailed analysis of the micrographs reveals a dependence of enzyme activity on the phospholipid organization and orientation in the bilayer. These experiments suggest that it is possible to observe single enzymes at work in small channels, which are created by the hydrolysis of membrane phospholipids. Indeed, the measured rate of hydrolysis of phospholipids corresponds very well with the enzyme activity found in kinetic studies. It was also possible to correlate the number of enzymes at the surface, as calculated from the binding constant to the number of starting points of the hydrolysis. In addition, the width of the channels was found to be comparable to the diameter of a single phospholipase A2 and thus further supports the single-enzyme hypothesis. 相似文献
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The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from
nucleotide sequence variation across a 765-bp region in the cytochrome
oxidase I and II genes of the mitochondrial genome. Most parsimonious
relationships of 25 haplotypes from 16 Greya species and two outgroup
genera (Tetragma and Prodoxus) showed substantial congruence with the
species relationships indicated by morphological variation. Differences
between mitochondrial and morphological trees were found primarily in the
positions of two species, G. variabilis and G. pectinifera, and in the
branching order of the three major species groups in the genus. Conflicts
between the data sets were examined by comparing levels of homoplasy in
characters supporting alternative hypotheses. The phylogeny of Greya
species suggests that host-plant association at the family level and larval
feeding mode are conservative characters. Transition/transversion ratios
estimated by reconstruction of nucleotide substitutions on the phylogeny
had a range of 2.0-9.3, when different subsets of the phylogeny were used.
The decline of this ratio with the increase in maximum sequence divergence
among taxa indicates that transitions are masked by transversions along
deeper internodes or long branches of the phylogeny. Among transitions,
substitutions of A-->G and T-->C outnumbered their reciprocal
substitutions by 2-6 times, presumably because of the approximately 4:1
(77%) A+T-bias in nucleotide base composition. Of all transversions,
73%-80% were A<-->T substitutions, 85% of which occurred at third
positions of codons; these estimates did not decrease with an increase in
maximum sequence divergence of taxa included in the analysis. The high
frequency of A<-->T substitutions is either a reflection or an
explanation of the 92% A+T bias at third codon positions.
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