We demonstrate that the sub-millisecond protein folding process referred to as "collapse" actually consists of at least two separate processes. We observe the UV fluorescence spectrum from naturally occurring tryptophans in three well-studied proteins, cytochrome c, apomyoglobin, and lysozyme, as a function of time in a microfluidic mixer with a dead time of approximately 20 mus. Single value decomposition of the time-dependent spectra reveal two separate processes: 1), a spectral shift which occurs within the mixing time; and 2), a fluorescence decay occurring between approximately 100 and 300 micros. We attribute the first process to hydrophobic collapse and the second process to the formation of the first native tertiary contacts.
Cell cycle and DNA repair involve steps requiring RanGTP-regulated nuclear-cytoplasmic transport (NCT). The activation of NCT via overexpression of RCC1 (the guanine nucleotide exchange factor for Ran) accelerated the cell cycle and DNA repair. Normal cells overexpressing RCC1 evaded DNA damage–induced senescence, mimicking cancer cells with high levels of RCC1.
Kinesin-5 mediates chromosome congression in Candida albicans via length-dependent depolymerase activity, which organizes chromosomes at the spindle equator to overcome fundamental thermal limits at the nanoscale and maintains spindle dimensions regardless of cell size.
Summary
Proper segregation of the replicated genome requires that kinetochores form and maintain bi-oriented, amphitelic attachments to microtubules from opposite spindle poles and eliminate erroneous, syntelic attachments to microtubules from the same spindle pole. Phosphorylation of kinetochore proteins destabilizes low-tension kinetochore-microtubule attachments, yet tension stabilizes bi-oriented attachments. This conundrum for forming high-tension amphitelic attachments is recognized as the “initiation problem of bi-orientation (IPBO).” A delay before kinetochore-microtubule detachment solves the IPBO, but it lacks a mechanistic framework. We developed a stochastic mathematical model for kinetochore-microtubule error correction in yeast that reveals: 1) under low chromatin tension, requiring a large number of phosphorylation events at multiple sites to achieve detachment provides the necessary delay, and 2) kinetochore-induced microtubule depolymerization generates tension in amphitelic, but not syntelic, attachments. With these requirements, the model provides a mechanistic framework for the delay before detachment to solve the IPBO and demonstrates the high degree of amphitely observed experimentally for wild-type spindles under optimal conditions.
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