Cells can mutate to express more or less AR.
Single or paired conserved residues were mutated to alanine, expressed in HEK293 cells, and lipid droplets were visualized by fluorescence confocal microscopy in order to characterize alterations in phenotype.
To investigate whether the corresponding sequence in UNG2 also constitutes a degron, UNG2 T60 and S64 were mutated to alanine, and expressed as EYFP fusions in HeLa.
PDI ΔS1S2 (a gift from Professor Lloyd Ruddock, Oulu Finland), in which both active sites are mutated to AGHA, was expressed and purified as described for the wt protein.
Residue W6.48 in the human adenosine A3 receptor fused to yellow fluorescent protein was mutated to phenylalanine and expressed in CHO-K1 cells containing a cAMP response element reporter gene.
To explore the contribution of the potential rim contacts to mediate the interaction between RydC and cfa, an Hfq mutant in the lateral rim (where residues R19, P21, Q33, Q35, T49, R66 have been mutated to A) was expressed and purified, and its binding rates and affinities were estimated from interferometry (ForteBio Octet Red96).
To address the technical challenges of expressing Plasmodium proteins recombinantly, codon-optimized expression constructs containing an exogenous signal peptide and in which N-linked glycosylation sequons were mutated to prevent inappropriate glycosylation were expressed in mammalian cells (Crosnier et al., 2011; Bartholdson et al., 2012).
Transgenic mouse models that solely express AβpE3 42 (Glu-3 mutated to Gln-3 in order to facilitate pyroGlu-3 formation) consistently develop neuron loss and associated neurological deficits.
Hence, these codons were mutated using oligonucleotide primers to express serine in the recombinant Hos2 protein.
Sequence analysis showed that most PERV proviral DNA was significantly mutated, thus suggesting the inability to express functional viral RNA; however, it cannot be ruled out that compensatory recombination processes could occur enabling replication of defective proviruses.
