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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5586_Библиотеки_им_академика_М_И_Перельмана.pdf
Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
• alignment-free trees based on statistical properties of the genome;
• gene content trees based on the presence and absence of genes;
• phylogenomics-based genome trees;
• trees based on average sequence similarity; and
• trees based on chromosomal gene order.
Despite their current advancement, genome tree approaches have already had some
influence on the phylogenetic arrangement of bacterial species. However, their main
impact so far has been on our understanding of the nature of genome evolution and
the role of horizontal gene transfer therein [57]. Relative genome analyses disclose
that most functional domains of human genes have homologs in commonly
divergent species. These shared functional domains, however, are differentially
shuffled among evolutionary lineages to create an increasing number of domain
architectures. Combined with duplication and adaptive evolution, domain shuffling
is accountable for the great phenotypic complexity of higher eukaryotes [83–85].
Mobile elements within genomes have determined genome evolution by different
means. Mainly in plants and mammals, retrotransposons have been shown to
establish a large fraction of the genome and have shaped both genes and the entire
genome. Although the host can frequently govern their numbers, massive expansions of retrotransposons have been accepted during evolution. Currently, mobile
elements are becoming valuable tools for learning more about genome evolution and
gene function.
Based on paleological investigation, it was demonstrated that approximately
3.5 billion years ago, cells identical to bacteria existed on our planet. Based on this
observation we can only accept that the genomes of these organisms or the
organisms that evolved soon afterward contain double-stranded DNA molecules.
According to previous findings (dated to about 1.4 billion years ago) the first
eukaryotic fossils were similar to single-celled algae, which means that both
prokaryotes and eukaryotes experienced variations in size, shape and complexity,
thus their genomes also transformed vigorously. These variations were motivated by
mutation, transposition, gene transfer and recombination, along with gene deletion
and duplication. Through investigating and equating genomes of organisms that
exist currently, we can gain understandings into how these mechanisms have shaped
genomes.
7.8.1 Microbial genome reduction in bacteria
Once bacterial pathogens make the evolution from free-living life cycles to a
permanent relationship with a host, they experience a significant loss of genes and
DNA [86]. Complete genome sequences offer information on how significant
genome reduction affects the developmental directions and metabolic capabilities
of obligate pathogens and symbionts [58]. Reconstruction of the gene content and
order of the last common ancestor of human pathogens can be achieved, for
example in the case of Mycobacterium leprae and Mycobacterium tuberculosis .An
understanding into which genes are the most indispensable for existence comes from
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equating the genomes of two related species of Mycobacterium: M. tuberculosis,
which is responsible for tuberculosis and M. leprae, which is the causative agent of
leprosy. As per reports, M. tuberculosis has 3959 protein-coding genes and a
4.41 Mb genome. In contrast, M. leprae has 1604 protein-coding genes and a
3.26 Mb genome. Therefore, during development or evolution, M. leprae has
misplaced or lost approximately 2000 genes (more than 50% of its ancestral gene
set). Several reasons can account for this, such as codon elimination or deletion and
deterioration, through mutation and incorporation of transposable elements, which
have disabled many genes. Consequently, metabolic functions vital for cell development have been abolished, and the bacterium develops very slowly; it multiplies once
every 14 days. Moreover, genetic mutations for recombination and DNA repair are
incapable of averting additional loss to the M. leprae genome. This blend of
mutations may have comdemmed M. leprae to extinction, as epidemiologists have
projected that the organism is at the limit of being able to sustain itself by infecting
new individuals [59].
7.8.2 Role of duplications in the origin and evolution of the eukaryotic genome
Assessments of prokaryotic and eukaryotic genomes are providing indications of the
lineages of the eukaryotic genome and by what means eukaryotes arose from
prokaryotes. Several lines of inquiry, comprising amino acid analysis of proteins,
gene sequences and metabolic pathways, determined that the eukaryotic genomes
are actually mosaics that have notable contributions from both the Archaea and the
Eubacteria. For example, the eukaryotic nuclear genome has few, if any, operons,
and its genes encompass introns, both of which are features of the Archaea.
The mitochondrial genome of a eukaryotic looks very much like that of alphaproteobacteria. To explain these observations, it has been suggested that eukaryotes
arose as a result of a symbiotic relationship, or fusion, between an anaerobic
archaebacterial host and an alpha proteobacterium (such as Rickettsia), which
developed into the mitochondrion [90]. The monophyletic character of all eukaryotes suggests that this incident occurred effectively only once in the history of the
Earth.
The duplication–divergence concept was suggested many decades ago, i.e., that
new genes evolve from pre-existing ones via gene duplication and subsequent
divergence of the extra copy to acquire a new function (figure 7.12). Based on this
concept Susumu Ohno wrote the book Evolution by Gene Duplication. Since then,
comparative genomics, genetics and biochemistry have clearly demonstrated that
duplication–divergence mechanisms are a key contributor to the evolution of new
genes. In contrast, the de novo origination model of new gene evolution is more
recent and less supported. De novo origination of protein-coding genes takes place
when genes arise from a nonfunctional DNA sequence that was formerly not a gene.
It would seem highly questionable whether functional proteins could arise spontaneously from non-coding DNA, as the DNA must both be transcriptionally active
and comprise a translatable open reading frame. Also, translation of any random
ORF devoid of genes is anticipated to offer unimportant polypeptides rather than
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Figure 7.12. Role of duplications in the origin and evolution of the eukaryotic genome.
proteins with specific functions. Certainly it has been claimed that de novo
origination of new genes is tremendously unlikely, however, irrespective of these
reports, the arrival of large-scale sequencing and comparative genomics has
delivered increasing proof that new genes have developed and continue to originate
from non-coding sequences [60].
Gene duplication plays a significant role in the development of eukaryotic
genomes, particularly their size and complexity. Susumo Ohno has predicted that
complete genome duplications are a powerful development mechanism [91].
Examination of nucleotide sequence information from genome projects encourages
the impression that a considerable portion of the difference in gene number that
differentiates prokaryotes from eukaryotes arose from genome expansions.
Currently, it is understood that a significant expansion in eukaryote genome size
arose from a genome duplication event that accompanied the arrival of vertebrates
in the fossil record. Genome duplications have also occurred at other times during
eukaryotic development. A current investigation of the yeast genome displays traces
of its earliest development by genomic duplication. The yeast genome has been
determined to hold at least 55 duplicated regions encompassing 376 genes, which
covers 5% of the genome. Of these 55 regions, 50 are in a similar comparative
location on diverse chromosomes. For example, chromosomes XI and XIII
comprise duplicated blocks of genes in which gene order and positioning relative
to the centromere has been well-maintained. Additional chromosomes cover internal
duplications, such as the region positioned on either side of the centromere on
chromosome XII. Phylogenetic examination designates that this genome duplication
incident took place around 100 million years ago. Examination of the human
genome also displays proof of an early large-scale duplication, tracked by reorganizations and gene damage in certain duplicated regions. These duplications range
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Figure 7.13. Functional genomics investigates the coordination among the genome, transcripts (genes),
proteins, and metabolites to generate specific phenotypes.
from minor sections covering only few genes to great stretches that cover nearly a
complete chromosome. The larger duplications date to the origin of the vertebrates,
around 500 million years ago. Overall, there are 1077 blocks of duplicated regions in
the human genome, covering 10 000 genes (around one-third of the genome). Such
duplication includes chromosomes 18 and 20. Different potential areas covered
under proteomics are presented in figure 7.13.
7.8.3 Gene duplications increase genetic diversity and complexity
Genome duplication is both a historic and continuing process in yeast, plants and
animals. It was recently established that the yeast genus Saccharomyces experienced
an ancient whole-genome duplication, and genome duplication is pervasive in
plants. Numerous incidents of genome duplication have occurred during the
divergence of angiosperms, including ancient polyploidization processes and the
assembly of self-governing duplications in different ancestries. Although not as
common as in plants, many genome duplications have also happened in independent
lineages in metazoans. During chordate development, whole-genome duplications
have occurred coincident with the origin of vertebrates gnathostomes, and teleosts.
Gene and genome duplications offer a basis of genetic material for mutation, drift
and selection to act upon, making new evolutionary changes more likely. Thus,
several researchers have claimed that genome duplication is a dominant factor in the
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evolution of complexity and diversity. However, a strong connection between a
genome duplication incident and increased complexity and diversity is not conclusive, and there are variations in the patterns of diversity raised to support this
claim. Remarkably, many studies of genome duplication processes in vertebrates
show they are preceeded by numerous extinct ancestries, resulting in preduplication
gaps in extant taxa.
Genomic nucleotide sequence information indicates that multigene families are
found in the human genome, if not all genomes. In addition to genome-wide
duplications, small chunks of genes and single genes can be duplicated by numerous
mechanisms, including:
• unequal crossing over and
• replication errors.
Throughout replication of a template molecule, a slippage can cause the introduction of a short segment into the freshly produced strand. When produced,
participants of multigene families may remain associated on a single chromosome
or can scatter to other parts of the genome. Numerous mechanisms determine this
procedure, such as inversions, translocations and transposition by mobile elements.
The ancestry and relationships among members of gene families have been
recently established by several groups of molecular phylogeneticists. Among the
most studied examples is the globin gene superfamily. Some 800 million years ago,
duplication of a family gene encoding an oxygen transport protein occurred in this
family. This has formed two sister genes, one of which developed into the modern
day myoglobin gene. Myoglobin is an oxygen-carrying muscular protein, i.e., it is
found in muscles. Around 500 million years ago, the ancestral globin gene
duplicated to produce the prototypes of the α- and β-globin subfamilies. The αand β-globin genes encode for the proteins that are present in hemoglobin, the
oxygen-carrying molecule in red blood cells. Further duplications within the α- and
β-globin genes occurred in the last 200 million years. Subsequent events dispersed
members of this superfamily, and each is now on a separate chromosome. Similar
patterns of development are reported in other gene families, including the trypsinchymotrypsin family of proteases, the homeotic selector genes of animals and the
rhodopsin family of visual pigments.
7.9 Algae bioinformatics
An alga is considered a unique source of various bioactive compounds and having
significant biological activities. Algae bioinformatics, as the name suggests is the
application of information technology to decipher more algae with the aid of
computational tools and software. This emerging field, just like the other streams of
bioinformatics employs computational or dry-lab techniques for applications in
various areas such as gene prediction, comparative genomics, genome analysis, and
functional genomics and so on. Algae bioinformatics cannot be performed without
computer science, biology and genetics with a good-sized dollop of mathematics,
statistics and other medical specialties thrown into the mix. There are many tools
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available for algae bioinformatics research, which are briefly discussed at the very
end of this article.
7.9.1 Scope of algae bioinformatics
Algae bioinformatics is a valuable resource for geneticists, phycologists and other
scientists who sequence algal genomes as a part of their wet-lab research. Therefore,
there is an immense requirement for more reliable, sophisticated, computerized
methods for examining this information bringing in the requirement of an algae
bioinformaticist.
7.9.2 What is involved in algae bioinformatics
Algae bioinformatics involves the development of novel algorithms and statistics
with which relationships among the different algal species can be measured.
Examination and elucidation of different types of data and their related information
is done. This data study involves DNA, RNA and protein sequences and structures.
Development of various techniques allows the efficient access and management of
different types of algal samples.
7.9.3 Role of algae bioinformatics
As discussed earlier, algae bioinformatics will only involve dry-lab research and it
requires the use of RNA, DNA, protein sequence data. A series of wet-lab work is
done so as to obtain the sequencing data and is subjected to bioinformatics analysis.
7.9.4 Steps involved in obtaining the data for analysis using bioinformatics
• Design of primers;
• Extraction of DNA, RNA;
• PCR amplification;
• Denaturizing gradient gel electrophoresis;
• Sequencing.
Nucleic acids are usually derived from the algal samples which are then utilized to
synthesize and design primers and then PCR amplification is done. As the products
of the PCR reactions are equivalent in their size, a denaturing gradient gel
electrophoresis is done to identify variation between the sequences of the products.
After this step algal bioinformatics work is performed. This can be achieved by
making phylogenetic trees, BLAST searches for similar sequences, and annotation
of the new sequences.
7.10 Functional genomics
Functioning genomics is an academic discipline that explores the relationship among
a living thing’s genetic composition, often termed the genome, and its discernible
characteristics, known as an organism’s phenotype. Recognizing the genetic
processes that determine the operational dynamics and interaction of genes inside
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an individual is the fundamental goal. The study of gene expression, gene function,
and research into the gene structure are all methods included in genomics with
function [61]. An overview of functional genomics is presented in figure 7.13.
7.10.1 Introduction to functional genomics
The branch of research known as operational genomics seeks to understand the
complex relationship that links the genome and its phenotype on a broad scale that
encompasses the whole genome. The strategies used in this work include utilizing
high throughput techniques to investigate genes’ activation and interacting patterns.
This facilitates the examination and comprehension of the actions performed by
these genes and the cellular processes that govern their roles inside an organism.
7.10.2 Transcriptomics: studying the RNA molecules
Transcriptomics relates to the science of the field as it examines a transcriptome,
which covers the totality of RNA molecules created by the genetic code during a
specific time. The procedure involves the examination of the expression of gene
designs, the recognition of transcription factors, and the exploration of functional
elements inside the genome. The sequencing of RNA is the technique that is often
used in the transcriptomics study. It gives us helpful information about how genes
are expressed and how the genome is structured [62].
7.10.3 Proteomics: understanding the world of proteins
Proteomics is an extensive examination of peptides on a broad scale, focusing
mainly on elucidating their functions and structural traits. The field of research
being discussed has significance in understanding changes in metabolism under
different conditions and assumes a pivotal role in the early detection of illnesses,
forecasting their results, designing medications, and monitoring disease progression.
Proteomics methods allow for in-depth analysis and characterization of its proteome
at various stages of development and physiology by measuring protein expression,
structure, function, interaction between molecules, and changes following translation [63].
7.10.4 Metabolomics: exploring cellular metabolites
Metabolomics is a contemporary field of study within the ‘omics’ sciences, focusing
on the comprehensive analysis of metabolites, including qualitative and quantitative
evaluations. These metabolites encompass essential intermediates and final products
that arise from metabolic processes. The integration of high-throughput analytical
techniques and bioinformatics enables the systematic identification and quantification of several metabolites, hence facilitating the comprehension of metabolic
disruptions and the elucidation of underlying mechanisms associated with diverse
diseases [64].
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7.10.5 Interactomics investigating protein–protein interactions
Interactomics, as a sub-field within the discipline of systems biology, is primarily
concerned with investigating protein–protein interactions and their consequential
impact on phenotypic traits. Various methodologies are utilized to investigate and
analyze the interactome, facilitating the comprehension of protein functionality and
regulation. The utilization of interactomics encompasses biochemical and clinical
domains, significantly contributing to understanding cellular signaling, disease
pathogenesis and advancing therapeutic approaches [65].
7.11 Structural genomics
The primary objective of structural genomics is to get comprehensive insights into
the spatial arrangement of all proteins encoded by genetic material. This discipline
extends beyond conventional molecular genetics and biochemistry. To offer a more
comprehensive comprehension of the genome. The main aims of this study are
comprehensive genetic and physical mapping and sequencing of the entire genome.
A crucial objective is elucidating the experimental structures of all potential protein
folds. Structural genomics holds promise in offering a comprehensive comprehension of several facets of cellular existence, encompassing metabolic activities, DNA
replication, transcriptional processes, protein synthesis, and protein folding [66]. The
flow of structural genomics is represented in figure 7.14.
Figure 7.14. Structural genomics, facilitated by the National Institutes of Health (NIH) Protein Structure
Initiative (PSI), established a collaborative network of research and resource centers. The primary objective
was to comprehensively explore the structural aspects of proteins, particularly in the uncharted territories of
the protein sequence space. A key challenge addressed by the PSI was elucidating the intricate connections
between protein sequences and their respective structures. The ultimate aim was to enhance accessibility to
structural data for the majority of proteins based on their gene sequences.
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7.11.1 Introduction to structural genomics
Structural genetics may be seen as an extension of genomics that encompasses the
domain that includes structural inquiry, aiming to uncover the features of genome
architecture. High-throughput methods are used to determine the structural features of
proteins across the whole genome. The insights gained by structural genomics are
valuable in many fields, including medication creation and a molecular-level under-
standing of illnesses, since they allow for modifying genes and DNA segments. One of
the main goals is to assign functional annotations to proteins whose activities have not
yet been established, mainly by analyzing their structural properties [67]. The aim may
be achieved using several methodologies, such as inferring distant homology relationships, identifying ligands, and characterizing electrostatic bands and cavities among a
protein’s structures. Genomic structural variants (SVs) are also studied in this field;
chromosomal rearrangements affect at least 50 bp. Understanding structural variations is of the highest relevance owing to their ability to give valuable insights into
mechanisms of evolution and the deep molecular foundations of illnesses such as
varied forms of carcinoma and neurodevelopmental conditions [68].
7.11.2 The approaches used in the domain of structural genomics
Deciphering a three-dimensional (3D) structure that composes biological macromolecules, having a significant focus on certain peptide regions, is the goal of
structural genomics. New methods have been created to find and analyze these
combinations, which helps us learn more about the molecular and functional settings
where these giant molecules work. The development of protein structure determination
pipelines is mainly attributable to structural genomics, which has led to significant
advances in identifying hitherto uncharacterized proteins. Notable advancements in
protein production and the rapid identification of new protein structures have resulted
from investigating proteins obtained from microbial dark matter and human diseases
[68]. Crystallography using x-rays, NMR (nuclear magnetic resonance) spectral
analysis, and cryo-electron microscopy (cryo-EM) are just a few examples of the
many methods included in structural genomics and are essential to the field’soverall
goals and objectives. These techniques are often supplemented by other methodologies, including connecting mass spectroscopy, small angle scattering of x-rays
(SAXS), and neutron diffraction. This collective approach offers a complete means of
investigating biological macromolecules. The challenges encountered within structural
genomics are manifold. The large volumes of data generated necessitate effective
collection, display, and analysis protocols to maximize the value derived from such
substantial investments [69]. Moreover, the quality of structures determined is a
concern, particularly when resolution data are artificially limited, which might
undermine the benefits of structural genomics endeavors.
7.11.3 Importance of structural genomics in drug design
Structural genomics holds significant promise for drug discovery. The growing
repository of high-resolution structures of known and potential drug target proteins
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is poised to benefit future drug discovery programs substantially. When the target
protein of a drug is known, structural genomics facilitates the rapid and efficient
acquisition of ligand-bound structures, leveraging high-throughput x-ray crystallography and NMR [70]. In cases where the drug target is unknown, structural genomics
provides purified proteins for numerous potential drug targets, thus supporting drug
discovery efforts. The impact is already being felt in the early stages of drug discovery
and target validation, with the contribution of new structures, complexes with ligands,
and supportive protocols and reagents for additional structural work within drug
discovery programs. The role of structural genomics in modern structure-based drug
design is underscored by its ability to analyze many target proteins concurrently.
While several structural genomics initiatives have been launched, a relative few have
focused on integral membrane proteins, which represent a critical area for future
exploration in drug design and discovery [71].
7.12 Epigenomics and epigenetics
Epigenomics is a sub-field of genomics that explores the comprehensive characterization and analysis of all epigenetic modifications across the genome. Epigenetics is
the study of reversible, heritable changes in gene function that occur without altering
the DNA sequence itself. DNA methylation is a hallmark epigenetic regulation
mechanism [72]. Epigenomics is concerned with understanding the broader epigenetic
landscape across the genome, focusing on modifications like DNA methylation,
histone modifications, and chromatin remodeling that do not alter the DNA sequence
but significantly impact gene expression, cellular function and phenotype. The
conventional concept of epigenetics, articulated initially by Conrad Waddington
during the 1950s, pertains to the investigation of enduringly inheritable phenotypic
traits that arise from modifications to a chromosome without concomitant alterations
to the DNA sequence [73]. Epigenetic alterations play a critical role in various
biological processes, such as development, differentiation, and the ability to respond to
environmental stimuli. The rapid acceleration of epigenetic research in the 21st century
has generated excitement and hope, linking genetics to environmental factors and
diseases [74]. DNA methylation, a pivotal epigenetic mechanism, involves adding a
methyl group to the cytosine base of DNA, predominantly at CpG dinucleotides.
DNA methylation patterns are established early in mammalian development and are
maintained during somatic cell division, playing crucial roles in gene silencing,
x-chromosome inactivation, genomic imprinting, and suppressing transposable element activity. These dynamic patterns can be altered in response to environmental
stimuli and during different developmental stages. For instance, the review highlights
the dynamic erasure and re-establishment of DNA methylation in embryonic, germline, and somatic cell development, emphasizing its significance in mice and humans.
DNA methylation and demethylation processes also occur in the nervous system,
associating with other epigenetic mechanisms like histone modifications and noncoding RNAs, showcasing the interplay between different epigenetic modifications
[75]. The structure and distribution of CpG islands and the role of methylation in gene
expression regulation, embryogenesis, ageing, and cancer development are also critical
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