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Hierachical clustering of the mutants with all measured parameters was performed to test whether there are groups of mutants with similar phenotypic pattern.

Previous studies showed that mutants with all 7 enzymes except for GA2ox exhibited a typical phenotype of dwarfism, indicating the functional importance of these enzymes in GA biosynthesis [ 23].

Eight NBD1/linker and eight NBD2 mutants that maintained the highest levels of resistance against all three fungicides (highlighted in Figure 5A) were chosen to be randomly assembled into mutants with all six intracellular Cys removed (C6Z).

HeLa cells stably expressing his6-SUMO2 were transiently transfected with plasmids expressing His6 -HA--tagged Kif18A (His6 -HA-Wtheor the mutant protein with single lysine residue replaced with arginine (K148R, K442R, K533R, K660R and K683R) or the mutants with all 5 lysine residues replaced with arginines (5R) for 48 h.

Within this group of mutations that modify the negative charge of SOD1, we can find SOD1 mutants with all defined levels of aggregation propensity (low, moderate, high or extreme), not following a clear correlation between a decrease or increase in protein charge and aggregation rates.

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To reveal the potential contribution from the phosphorylation of the three BubR1 KARD domain residues, we generated a BubR1 mutant with all three sites mutated to aspartic acid (S670D/S676D/T680D), which was previously used to mimic BubR1 phosphorylation (Suijkerbuijk et al., 2012).

The Gap1K9,16R mutant was still down-regulated in this mutant context and only a Gap1 mutant with all nine lysines present in NT replaced with arginine turned out to be protected against degradation and stabilized at the plasma membrane [5].

In this vein, we have characterized a zebrafish mutant with all of the relevant features of NM.

Similar to rad8 +, the helicase-dead mutant rad8-K535A, T536A (rad8-HD), ubiquitin ligase-dead mutant rad8-I879A (rad8-LD), and the mutant with all three mutations rad8-K535AT536AI879A (rad8-HDLD) were all nuclear localized.

Cells co-expressing NlpΔ8 mutant (with all eight plk1 phospho-sites mutated to alanine) and Plk1 T210D are tremendously resistant to fragmentation of centrosomal Nlp into cytoplasm, which in turn contributes to false spindle formation.

The fact that all mutants shift with all tested proteins indicates they are correctly folded, however with our assay it is difficult to detect subtle differences in the affinity of the mutants for the ABPs (see also (11)).

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