Correcting Errors attachment

Figure 5.18 shows the only reliable Nui c data available near the critical Reynolds number (XI). Since the data were taken with a side support, there is some effect on the separation and transition angles. Thus the values of Nuj are probably subject to error (R2, R3) although the trend with Re should be correct. At Re = 0.87 x 10 the Shi variation is similar to that shown at lower Re in Fig. 5.17. At Re = 1.76 x 10 the critical transition has already occurred, with the separation bubble accounting for the minimum in Nuj c at 0 — 110°. The maximum in Nuj at 0 = 125° reflects the increased transfer rate in the attached turbulent boundary layer. The local minimum at 0 = 160° is due to final separation. These angles do not agree exactly with those in Fig. 5.11 because of the crossflow support and the fact that angular diffusion shifts the...

When action level excursions or frequent alert excursions are identified, a corrective action program, resolution deadline, and preventive plan shall be implemented. Risk analysis shall be performed to determine the probability of one or more causes of errors occurring, as well as to identify the potential consequences of excursions. (See attachment nos. 1700.80(1), 1700.80(J), and 1700.80(K), and 1700.80(L) for excursion of air, surface, personnel, and visual inspection report during the visit to plant by the microbiologist responsible.)... 

Synthases Attach Amino Acids to tRNAs Each Synthase Recognizes a Specific Amino Acid and Specific Regions on Its Cognate tRNA Aminoacyl-tRNA Synthases Can Correct Acylation Errors... 

Error correction is thought to occur by stabilizing correct attachments while destabilizing incorrect attachments (41). Experiments in yeast showed that the inhibition of the Ipll/Aurora family of kinases prevents error correction by stabilizing incorrect attachments (38, 42), but how the active kinase corrected attachment errors was not known. This problem was particularly difficult to address because attachment errors are observed infrequently in the presence of active Aurora kinase (43). Experimental approaches that accumulated attachment errors through inhibition of Aurora kinase, for example by genetic mutation (42), did not permit subsequent kinase activation to examine error correction. Reversible small-molecule Aurora kinase inhibitors present a solution to this problem because they can be used to inhibit kinase function and subsequently removed to activate the kinase. 

To devise a strategy to address the question of how attachment errors were corrected, several issues needed to be addressed. Eirst, Aurora kinases have been implicated in multiple processes in mitosis (44). Ideally, kinase inhibition temporally would be controlled to isolate experimentally the error correction process. Second, the microtubules fibers that attach chromosomes to the spindle are highly dynamic, and the error correction likely involves some regulation of these dynamics. Live imaging would permit the analysis of microtubule dynamics with high temporal and spatial resolution. Finally, analysis of microtubule dynamics is difficult if individual fibers are obscured by other microtubules in the spindle. By creating conditions in which the improperly attached chromosomes are positioned away from the spindle body, individual fibers could be observed clearly. 

Figure 4 Correction of improper chromosome attachments by activation of Aurora kinase (45). (a) Assay schematic, (i) Treatment with the Eg5 inhibitor monastrol arrests cells in mitosis with monopolar spindles, in which sister chromosomes often are both attached to the single spindle pole, (ii) Hesperadin, an Aurora kinase inhibitor, is added as monastrol is removed. As the spindle bipolarizes with Aurora kinase inhibited, attachment errors fail to correct so that some sister chromosomes are still attached to the same pole of the bipolar spindle, (iii) Removal of hesperadin activates Aurora kinase. Incorrect attachments are destabilized by disassembling the microtubule fibers, which pulls the chromosomes to the pole, whereas correct attachments are stable, (iv) Chromosomes move from the pole to the center of the spindle as correct attachments form, (b) Structures of the Eg5 inhibitor monastrol and two Aurora kinase inhibitors, hesperadin and AKI-1. (c) Spindles were fixed after bipolarization either in the absence (i) or presence (ii) of an Aurora kinase inhibitor. Arrows indicate sister chromosomes that are both attached to the same spindle pole. Projections of multiple image planes are shown, with optical sections of boxed regions (1 and 2) to highlight attachment errors. Scale bars 5 xm. (d) After the removal of hesperadin, GFP-tubulin (top) and chromosomes (bottom) were imaged live by three-dimensional confocal fluorescence microcopy and DIC, respectively. Arrow and arrowhead show two chromosomes that move to the spindle pole (marked by circle in DIC images) as the associated kinetochore-microtubule fibers shorten and that then move to the center of the spindle. Time (minutes seconds) after the removal of hesperadin. Scale bar 5 (cm.

Although the accuracy of translation (approximately one error per 104 amino acids incorporated) is lower than those of DNA replication and transcription, it is remarkably higher than one would expect of such a complex process. The principal reasons for the accuracy with which amino acids are incorporated into polypeptides include codon-anticodon base pairing and the mechanism by which amino acids are attached to their cognate tRNAs. The attachment of amino acids to tRNAs, considered the first step in protein synthesis, is catalyzed by a group of enzymes called the aminoacyl-tRNA synthetases. The precision with which these enzymes esterify each specific amino acid to the correct tRNA is now believed to be so important for accurate translation that their functioning has been referred to collectively as the second genetic code. 

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