We show experimentally that increased expression of the early APC/CCdc20 substrate Clb5 does not delay the degradation of the later substrate securin, arguing against a role for competition with Clb5 in establishing securin degradation timing.
Later, all substrates underwent an Au deposition and annealing step, so as to form isolated Au nanoparticles.
Later, complex substrates such as starch, cellulose, dextran, molasses, chitin and pectin (Niessen et al. 2005, 2006; Rezaei et al. 2007) were also employed.
At later times substrate depletion and the instability of all 3 proteases leads to lower rates of substrate turnover.
Later, the substrate relationship became less evident, or present only in very early developmental stages, with adult specimens becoming free-living forms, such as observed within Flabellum.
Then streptavidin peroxidase and later the substrate solution were added to detect the products of the reaction.
Later, each substrate was sieved (mesh size of sieves: 250 μm and 37 μm) under running water; then animals were picked up from the sieved sediments with a glass pipette under a stereomicroscope.
Later, the two substrates were well aligned under a microscope with the assistance of markers on both substrates and were eventually brought into contact to form permanent bonding.
For this particular experiment, we used the silica substrates (later, for demonstration purposes in the "Discussion" section, also silicon and oxidized silicon substrates were also used).
Therefore, we can explain the lower degree of conservation by the emergence of chaperonin-dependent substrates later on in evolutionary history.
