Diversity gene modifications

The key feature of the PDA technology is that a far wider biological diversity can be screened computationally than it is possible with exclusively experimental methods. The in silico approach provides unique and original gene modifications whilst maintaining the precise control over the extent and nature of protein modifications. The novel sequences... 

There is a broad diversity of heterologous hosts for gene expression, and there is no best or universal host which is suitable for the production of all possible proteins. Nevertheless, many proteins can be expressed cheaply and successfully in Escherichia coli, which is still the most preferred host for heterologous production of proteins without posttranslational modifications. 

Figure 11.5 Amino acid building blocks are incorporated into daptomycin backbone successively by NRPS subunits DptA, DptBC and DptD (a). Structural diversity of daptomycin peptide core can be obtained by genetic modifications of dpt gene cluster (b). C, condensation domain A, adenylation domain PCP, peptidyl carrier protein E, epimerase TE, thioesterase domain... 

The activity of PK and NRPSs is often precluded and/or followed by actions upon the natural products by modifying enzymes. There exists a first level of diversity in which the monomers for respective synthases must be created. For instance, in the case of many NRPs, noncanonical amino acids must be biosynthesized by a series of enzymes found within the biosynthetic gene cluster in order for the peptides to be available for elongation by the NRPS. A second level of molecular diversity comes into play via post-synthase modification. Examples of these activities include macrocyclization, heterocyclization, aromatization, methylation, oxidation, reduction, halogenation, and glycosylation. Finally, a third level of diversity can occur in which molecules from disparate secondary metabolic pathways may interact, such as the modification of a natural product by an isoprenoid oligomer. Here, we will cover only a small subsection of... 

Expression analysis using DNA microarrays analyzes only the transcriptome it should be mentioned that mRNA abundance in a cell often correlates poorly with the amount of protein synthesized (27). Important regulation takes place at the levels of translation and enzymatic activities. The only effect of a signal transduction pathway that is observed in a gene expression experiment is that at the endpoint of a given pathway. DNA microarrays currently have little value in determining post-translational modifications, which influence the diversity, affinity, function, cellular abundance, and transport of proteins. 

Compared to the genome, the proteome (the entire diverse protein content of a cell) is a far more dynamic system. Proteins imdergo post-translational modifications such as phosphorylation, glycosylation and sulphation, as well as cleavage for specific proteins. These alterations determine protein activity, localisation and turnover. All are subject to change following a toxic insult and, in some ways, the study of proteins holds more promise than the study of gene expression as the former is nearer to key activities in the cell. 

All classes of RNA transcripts must be processed into mature species. The reactions include several types Nucleolytic cleavage, as in the separation of the mature rRNA species from the primary transcript of RNA polymerase I action Chain extension (non-template-directed), as in the synthesis or regeneration of the common CCA sequence at the 3 end of transfer RNAs or of PolyA at the 3 end of mRNAs and Nucleotide modification, for example, the synthesis of methylated nucleotides in tRNA or rRNA. These reactions are a feature of both prokaryotic and eukaryotic gene expression, and the biological consequences are diverse. For example, modified nucleotides can affect the way in which a tRNA recognizes different codons. 

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