Fundamentals of Bioinformatics - Ogurtsov, A.N. 2013

Information Principles in Biotechnology
Genome Analysis
Genome Composition

METABOLISM/28.html">The Genetic Code exhibits a relatively high level of noise Immunity. Therefore, having complete genome information at hand, It is interesting to explore the evolutionary rates of Mutations by distinguishing between synonymous nucleotide substitutions—codon changes that do not alter the encoded amino acid—and non-synonymous substitutions, which result in mutations within the encoded protein (missense mutations).

Given two aligned Gene sequences, we can calculate K_a, the number of non-synonymous substitutions, and K_s, the number of synonymous substitutions. A high K_a/K_s ratio indicates positive Selection, which may be associated with functional divergence.

Comparative Genomics utilizing sequenced genomes enables the investigation of the following issues.

✵ Which genes account for phenotypic differences among individuals? Which genes are unique to each specific individual phenotype? Does their genomic Location vary from one individual to another?

✵ Which homologous Proteins account for phenotypic differences among individuals? Which proteins are unique to each specific individual phenotype? Does the cumulative effect of these proteins vary among individuals? Do The regulatory mechanisms controlling the normal expression of these proteins change?

✵ Which biochemical Functions account for phenotypic differences among individuals? Which biochemical functions are unique to each specific individual phenotype? Does the cumulative effect of these biochemical functions vary from one individual to another? Furthermore, if two individuals share a certain function, such that one possesses the protein responsible for it while the other possesses its homolog, does this protein perform the exact same function In the second Organism?

Currently, the following major functional classes of proteins are widely accepted:

✵ Cell energetics.

- Biosynthesis of Cofactors and Amino Acids.

- Central and Intermediary Metabolism.

- Energy Metabolism.

- Fatty acids and Phospholipids.

- Nucleotide biosynthesis.

- Transport.

✵ Information.

- Replication.

- Transcription.

- Translation.

✵ Communication and regulation.

- Regulatory functions.

- Cell envelope / Cell wall.

- Cellular processes.

When comparing the protein sets of three distinct species—the bacterium Haemophilus influenzae (1,680 genes), the archaeon Methanococcus jannaschii (1,735 genes), and the eukaryote Saccharomyces cerevisiae (Yeast, 6,278 genes)—researchers investigated whether these organisms share common proteins for common functions.

Within The Cell energetics Class, proteins performing energetic functions are evenly distributed across all three species. In the communication class, proteins are unique to each species. In the regulation and information classes, archaea share certain proteins with Bacteria, while sharing others with eukaryotes.

Genome comparison allows us to approach The Challenge of creating a so-called "minimal organism," that is, an organism with a minimal genome consistent with the Central dogma of molecular biology, DNA→RNA→protein (a dogma that rejects the existence of protein-free life forms based solely on RNA). A minimal organism must possess the capacity for self-replication, though it does not necessarily need to compete with other organisms in terms of GROWTH AND REPRODUCTION rates. It can be hypothesized that a minimal organism must be capable of assimilating a nutrient medium to sustain biosynthesis, as well as mounting a stress response, which includes the Repair of Damaged DNA.

The genes that such a minimal organism must contain are called operational genes or housekeeping genes. These are the genes necessary to maintain the vital functions of an organism, which are expressed in virtually all Tissues and Cells at a relatively constant level. Housekeeping genes function universally across all stages of an organism's life cycle.

Analysis of sequenced genomes has shown that the minimal genome should include the following functional classes of genes.

✵ Translation and Protein Synthesis.

✵ DNA replication.

✵ DNA recombination and repair.

✵ Transcription apparatus.

✵ Chaperone proteins.

✵ Intermediary metabolism - The Glycolytic Pathway.

✵ Structures ensuring transmembrane protein translocation.

✵ An essential set of transport proteins responsible for delivering missing metabolites from the environment.

However, it has not yet been proven that this necessary set of proteins is sufficient to ensure the viability of an organism. For example, a comparison of eukaryotic and archaeal genomes revealed only 71% of homologs from the proposed set of 256 proteins, and there is no reason to assume that the remaining 29% lack genes vital for the functioning of a minimal organism.

Nevertheless, functional definition is undoubtedly a universal approach in studying all forms of life, allowing us to investigate the extent to which different organisms have shaped these functions through parallel evolution. For instance, whether Homologous proteins from different species catalyze similar reactions.

Genome Analysis has revealed families of homologous proteins in archaea, bacteria, and eukaryotes.

Evolutionary bioinformatics makes it possible to begin testing many hypotheses that are widely accepted today.

The first such hypothesis is that Protein Families evolved from an individual ancestral gene through speciation and duplication events.

An alternative to this is Horizontal Gene Transfer—a process in which an organism transfers genetic material to another organism that is not its descendant (unlike horizontal transfer, vertical gene transfer occurs when an organism receives genetic material from its ancestor).

The ultimate goal of this kind of informational analysis is to map out the common functions and shared Proteins of the compared organism classes, which is currently a subject of intensive research in the field of evolutionary bioinformatics.

Several thousand protein families have already been established through Homology analysis with archaea, bacteria, and eukaryotes. Different species contain varying numbers of these shared families: the bacterium Aquifex aeolicus has 83% of its proteins with homologs among archaea and eukaryotes, whereas Borrelia burgdorferi has only 52%. Archaeal genomes possess a slightly higher percentage of proteins (62–71%) homologous to bacterial and eukaryotic proteins. However, only 35% of yeast proteins are homologous to bacterial and archaeal proteins.

Another widely accepted hypothesis is that a common set of proteins performs a common set of functions.

Among the minimal set of proteins identified in M. genitalium, only about 30% show homology across all known genomes. Other essential functions must be carried out by non-homologous proteins or, in some cases, by unrecognized homologs.

The total number of protein families in which homologs perform shared functions in archaea, bacteria, and eukaryotes increases significantly due to proteins involved in Translation Processes (Table 18).

Thus, the Analysis of Protein family evolution during the adaptation of proteins to perform life-critical functions indicates that such proteins should be the most conserved among the entire array of proteins synthesized by organisms.

Table 18 - Number of protein families characteristic of all known genomes

A comparison of two groups of yeast genes—those inherited from archaeal ancestors versus those from eubacterial ancestors—revealed that, by all measures, genes of archaeal ancestry are more critical for yeast cell viability. James A. Cotton and James O. McInerney demonstrated that the higher importance of archaeal genes does not depend on their function: both informational genes (involved in transferring information from DNA to proteins) and operational genes (housekeeping genes) of archaeal origin are more vital to the cell than those of eubacterial origin.

A database of yeast gene deletions (or knockouts) was utilized. This database contains information on gene functions elucidated by disrupting specific genes. By removing a gene from The Genome, researchers can observe whether cells can survive without it. If cells lacking the eliminated gene die, the deletion is classified as lethal; if they continue to reproduce normally, the deletion is deemed non-lethal. Geneticists divided the genes corresponding to lethal and non-lethal yeast deletions into two groups: those of archaeal origin and those of eubacterial origin. Naturally, a third group comprised unique eukaryotic genes, though they were not the focus of this study.

This seemingly simple Classification yielded unexpected results. Among the yeast genes, archaeal deletions proved to be 2.5 times more likely to be lethal than eubacterial ones. Furthermore, the expression level of archaeal genes was more than double that of eubacterial genes.

Of course, it is well established that the roles of archaeal and eubacterial genes are fundamentally different.

The archaeal gene group is primarily involved in information Processing—gene translation, transcription, and replication—whereas the eubacterial group is responsible for cellular metabolism. Could the differences in deletion lethality and expression levels be attributed to this functional significance for cell viability? If a metabolic pathway is disrupted, the cell can quite possibly utilize an alternative bypass route or manage without the final synthesis product. However, if a link in the information-processing machinery is disabled, the cell will inevitably perish.

Nevertheless, the differences in lethality between archaeal and eubacterial deletions do not depend on the gene's functional domain. Archaeal genes involved in Metabolic pathways are still, on average, more critical than their eubacterial counterparts; conversely, eubacterial participants in information processing are less vital for life support than archaeal representatives.

Thus, it became clear that archaeal genes play a more vital role in yeast cells than eubacterial ones, despite being fewer in number. A gene's evolutionary Lineage turns out to be just as significant in explaining its vital importance (lethality), interactions with other genes, expression level, and other parameters as the functions it performs.

Evolution is forced not only to address current environmental demands but also to carry the historical baggage of past adaptations.

Cells and organisms must sustain the thread of life that extends to them from the earliest primordial cells. Evolution is constrained by the necessity of passing on acquired refinements through successive generations. Once a critically important machinery has been perfected and is in continuous operation, it becomes exceedingly difficult to remove, replace, or attempt to modify it, as doing so carries a very high risk of disrupting the entire system—resulting in cell death.

This holds particularly true for such key molecular processes as

✵ reading and utilizing Genetic information;

✵ generating energy essential for cellular function;

✵ reproduction.

Each of these processes depends critically on the coordinated operation of dozens and hundreds of molecular machines.

This has led to a remarkable standardization, uniformity, and similarity in how such core processes operate at THE MOLECULAR LEVEL across all biological systems. They are all constructed from similar fundamental components discovered by evolution long ago and utilized ever since in all subsequent organisms.