imagine (Institute of molecular genetics and genetic engineering)
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Two contrasting late embryogenesis abounded protein family groups of Ramonda serbica Panc.
Get PDFRamonda serbica Panc. is an ancient resurrection plant, that survives a long desiccation period
and fully recovers metabolic functions upon watering. The main characteristic of desiccationtolerant
plant species is their ability to accumulate protective late embryogenesis abounded
protein (LEAPs). To propose their role in R. serbica desiccation tolerance we structurally
analysed LEAPs in hydrated and desiccated leaves.
According to transcriptomics, 318 LEAPs were identified and classified into seven family
groups based on protein BLAST analysis and conserved motifs (Pfam). The largest LEAPs
belonged to the LEA2 and LEA4 protein family groups. We employed online tools to analyse
physicochemical characteristics (Expasy, ProtParam, BioPython, GRAVY calculator), disorder
propensity, and characterization protein structures (FELLS, JPred, SOPMA, PsiPred, Phyre2,
Espritz-DisProt, Espritz-X, Iupred, TMHMM, +Heliquest).
The most abundant, atypical LEA2 group containing 127, mostly hydrophobic proteins, was
divided into five subgroups. Members of this group were predicted to fold into globular
domains, β-barrel at the C-terminus, followed by transmembrane hydrophobic-helices and
disordered N-terminal regions. Results indicated the possible involvement in the protection
of the chloroplastic membranes.
The LEA4 group exhibited an exceptionally high tendency to form amphipathic α-helices and
simultaneously had a high disorder propensity. This group is made of 96 proteins, classified into 3
subgroups. The high content of polar and charged amino acids (lysine, glutamate, and aspartate) is
characteristic of this group. Motifs corresponding to the R. serbica LEA4 protein family group folded
into A-type α-helices that contained positive, negative, and hydrophobic surfaces. Based on previous
knowledge, the possible functions of the LEA2 and LEA4 groups are discussed with significant
implications on cell preservation technology and the improvement of crop drought tolerance.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202
Plasma assisted bio-degradation of poly-lactic acid (PLA)
Get PDFPlastics are artificial synthetic organic polymers that have been used in every area of daily life. However, because of their slow degradation rate, their use
is contentious. The treatment of the surface of the sample is considered necessary as enzymatic or bacterial attach is not possible, if the plastic surface
environment is not ideal. The main topic of this work is the investigation of the effect of atmospheric dielectric barrier discharge (DBD) plasma on the near
surface structure of polylactic acid (PLA) samples, which, in turn, can promote the adhesion of enzymes or bacteria for further biodegradation. In general,
plasma processes can already be considered as inherently environmental technologies.
Plasma processes enable resource saving through high energy utilization efficiency and thus, are environ-mentally friendly technologies. Atmospheric
pressure discharges (APDs) are useful because of their specific advantages over low-pressure ones. They do not need expensive vacuum equipment, and
generate nonthermal plasmas, which are more suitable for assembly line processes. Hence, this category of discharges has significant industrial
applications. The use of a dielectric barrier in the discharge gap helps prevent spark formation. DBDs exhibit two major discharge modes: filamentary and
glow (homogeneous). The glow discharge mode has obvious advantages over the filamentary one for applications such as treatment of surfaces and
deposition of thin films. Glow mode discharges with average power densities comparable to those of filamentary discharges are of enormous interest for
applications in which reliable control is required.
Here we will present the increased adhesion of bacteria strains on DBD plasma treated PLA foils which can lead to a better degradation of the PLA. X-ray
photoelectron spectroscopy (XPS) measurements of the foils prior to and after the treatment proved the changes on the polymer surface. A short
discussion of the possibilities the treatment opens is given.CHANIA 2023: 10th International Conference on Sustainable Solid Waste Management Chania, Greece, 21 - 24 JUNE 202
In silico pre-selection of β-glucosidase gene for heterologous recombinant expression
Get PDFBiofilms are ubiquitous in nature, and the food industry is vulnerable to the risks posed by
biofilm formation. Not only do they interfere with the food production process, but they also
pose a public health threat. However, complete elimination of biofilms on food and food
contact surfaces cannot be achieved by conventional methods (cleaning and disinfection)
alone. New biofilm control strategies must be developed to prevent its formation and/or
persistence. Novel approaches may be based on enzymes that depolymerize components
of the biofilm matrix, making bacterial cells accessible to antimicrobial agents.
Environmental microorganisms are an inexhaustible source of new enzymes. In
Salmonella Enteritidis and Escherichia coli, known foodborne pathogens, cellulose is an
important component of the biofilm matrix, so our isolates from untapped environments
were tested for cellulolytic activity. Of the more than 70 isolates examined, isolate BG28
was selected as the most promising. Its genome was sequenced, annotated, and it was
identified as Gram-positive Microbacterium sp. Genome mining revealed the presence of
four complete genes for different β-glucosidases, one of three enzyme types of cellulase
complexes. To select the best candidate for heterologous expression DeepTMHMM,
ProtParam, and SoluProt were used to predict the presence/absence of signal peptide
and transmembrane domains, instability index, aliphatic index, hydrophilicity, and soluble
expression in E. coli. Based on the prediction results, the gene annotated as β-glucosidase
B was selected for recombinant expression. In addition, I-TASSER was used to model the
tertiary structure of the selected enzyme.
The β-glucosidase B was recombinantly expressed, purified, and tested for its anti-biofilm
activity. It was active and showed a 50% inhibitory effect on S. Enteritidis and E. coli biofilm
formation at a concentration of 100 μg/ml. To further evaluate this in silico approach in
the preselection of candidate enzymes for recombinant expression and purification, we
will use it to identify other enzymes of the cellulase complex.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202
Circular Codes in the Genetic Information
Get PDFCodes are the sets of words over arbitrary alphabets with the property of unique
decipherability.
Circular codes are a special class of codes. They are the sets of words with the property
of unique recognition of the reading frame for any sequence composed of them and
written on a circle. They were introduced by Golomb and Gordon in the 60s under the
name of codes with bounded synchronization delay, because they have a strong property
of synchronization. For this reason, they play an important role in problems of error
correction.
In the middle 90’s such a circular code X was identified in the genes of bacteria, eukaryotes,
plasmids, and viruses by a comprehensive statistical investigation. The code X contained
the 20 trinucleotides that appeared to be the codons that had the highest preference for
the correct reading frame compared to frames 1 and 2. Since then intensive research
on circular codes in the genetic information and their potential role in maintaining the
correct reading frame during the translation process in the ribosome has been done by
various authors. In particular, X-motifs were identified in (i) genes “universally” (ii) tRNAs
of prokaryotes and eukaryotes; (iii) rRNAs of prokaryotes (16S) and eukaryotes (18S), in
particular in the ribosome decoding center where the universally conserved nucleotides
G530, A1492, and A1493 are included in the X-motif; and (iv) genomes (non-coding
regions of eukaryotes). Circular codes have a highly complex structure and the ones found
in genes possess additional properties like e.g. self-complementarity that reflect their
biological nature.
In our talk we give a short introduction to the theory of circular codes and an overview on
the methods from mathematics, statistics and bioinformatics to explore their properties
and their biological role. Finally, a possible model of the evolution of the genetic code from
the perspective of circular code theory is presented.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202
Transcriptome analysis of Atdss1 mutants in response to oxidative stress
Get PDFDSS1 (deletion of split hand/split foot 1) is a highly conserved, eukaryotic, and multifunctional
protein. DSS1 as a small intrinsically disordered protein binds to multiple proteins when it
gains a final conformation. There are two highly homologous genes, DSS1(I) and DSS1(V) in the
Arabidopsis genome. Our aim is to examine Atdss1 mutants through oxidative stress. We
obtained separate stable lines of Arabidopsis containing mutations in DSS1s using
CRISPR/Cas9 technology. After H2O2 treatment, mutant seedlings showed increased
sensitivity to oxidative stress in comparison to WT plants. Transcriptome analysis showed
that dss1(I)del25 and dss1(V)ins18 mutations caused 2762 and 2335 differentially expressed
genes compared to WT under oxidative stress, respectively. We found that upregulated
expression was in genes involved in homologue recombination and RNA transport in both dss1
lines. The most downregulated genes are classified into flavonoid biosynthesis and MAPK
signaling pathway
Analysis of transcripts from alternative PRKAR1B gene promoters in colorectal cancer
Get PDFBackground/Objectives: The transcriptional regulation of
PRKAR1B is controlled by alternative promoters, and previous in
silico analysis has indicated their differential activity in colon and
rectal cancer tissue in comparison to normal gut mucosa. The aim
of this study was to investigate PRKAR1B promoters and transcripts
potentially involved in cancer.
Methods: The sequences of PRKAR1B alternative promoters
were retrieved from Ensembl database: promoter A 752209 and
promoter B 767287 bases upstream from the translation start site.
Bioinformatic tools Alggen, AliBaba, CiiiDER, and TFBIND were
used to predict binding of transcriptional regulators. Primer
extension assay was performed on RNA isolated from malignant
colon cell lines using an oligonucleotide probe binding to the
sequence at the exon2/exon3 junction common for all PRKAR1B
transcripts.
Results: Based on analyzed elements, both PRKAR1B promoters
were found to have atypical structure. According to the prediction,
promoter A that encodes transcript PRKAR1B-201 binds several
factors involved in cell proliferation, while promoter B that encodes
transcript PRKAR1B-203 binds mostly pro-apoptotic factors. In primer
extension experiments, a single signal corresponding to the
transcript PRKAR1B-212 was observed in malignant cells.
Conclusion: The differential activity of alternative PRKAR1B
promoters in colorectal cancer can be explained by in silico
results, predicting that promoter sequences bind sets of transcriptional
regulators with opposing roles. However, experiments
point to the transcript unrelated to either of the investigated
promoters as potential cancer biomarker and it should be further
characterized.Abstracts from the 55th European Society of Human Genetics (ESHG) Conference: e-Posters; Vienna, Austria.
June 11–14, 202
Echocardiography-based Left Ventricle Cardiac Hypertrophy Simulations
Get PDFClinical scenarios can be evaluated using numerical modeling of the cardiac cycle prior to
experimental or clinical application. Changes in wall thickness, displacement fields, and general
cardiac function are all affected by hypertrophy. In our study, we calculated the effects of
eccentric and concentric hypertrophy and monitored changes in ventricular thickness and shape.
Concentric hypertrophy results in thicker walls, while eccentric hypertrophy results in thinner
walls. Passive stresses were calculated using recently established material modals based on
Holzapfel’s work. Our modeling approach is based on composite shell finite elements, allowing
easier and more efficient modeling compared to traditional 3D finite elements. A left ventricular
model was constructed using echocardiographic images. Our modeling technology is based on
accurate patient-specific geometries and realistic constitutive curves, so it can be used as the
basis for real-world applications. Our model can be used to test medical hypotheses about the
development of hypertrophy in healthy and diseased hearts under the influence of different
conditions and factors.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202
Maqui berry extract inhibits filamentation of Candidaalbicans and improves the antifungal efficacy of nystatin
Get PDFCandidiasis caused by Candida albicans is one of the most common fungal infections in modern society. The limited arsenal of clinical drugs, their side effects and emerging resistance, largely contribute to the low efficacy of current antifungal therapies. The morphogenetic yeast-to-hyphae transition is the key virulence feature for the establishment of local and systemic C. albicans infections. We show that a delfinidin-rich fruit extract from Aristotelia chilensis ([Molina], Stuntz) (maqui berry) inhibits filamentation of C. albicans, in both laboratory and clinical strains. The extract acts synergistically with nystatin, with filament formation completely prevented by the combination of ¼×MIC of nystatin and 0.125 mg/ml of the extract. The combination treatment results in increased survival of C. albicans-infected zebrafish embryos compared to treatment with nystatin. Neither the extract nor its combination with nystatin was toxic at effective doses. These results warrant further investigation of maqui berry extract as adjuvant antifungal treatment
Evaluating ND1 and Cytb mitochondrial genes as markers for diversity analysis of protected White-tailed eagle species from Serbia
Get PDFWhite-tailed eagle is the biggest bird of prey in Central and Southeast Europe. In Serbia
it inhabits the Vojvodina province and the valleys of Danube, Sava, Tisa and Tamiš.
Anthropogenic pressure on its habitats in Europe caused a decline in its numbers, but
due to the strict laws protecting both species and its habitats, birds’ numbers are now
steady and increasing. In Serbia, as a strictly protected species it is a subject of different
conservation programs. The available genetic data for this population are scarce and it
is necessary to assess its genetic diversity to improve the existing conservation efforts.
ND1 and Cytb mitochondrial genes can be used to estimate the populations’ adaptation to
different environmental conditions and their variability can potentially be used to evaluate
differentiation between populations.
To assess the genetic diversity of White-tailed eagle in Serbia we used mitochondrial ND1
and Cytb nucleotide sequences from 40 unrelated birds collected in nests. ND1 and Cytb
nucleotide sequences variability was evaluated using standard parameters of genetic
diversity (PGD). Acquired values were compared with the available data for the variability
of the D-loop region which showed that combined ND1/Cytb nucleotide sequences PGD
provide comparable results. Using publicly available sequences we reconstructed haplotype
networks for ND1, Cytb, ND1/Cytb and D-loop which further showed the applicability of ND1/
Cytb in population genetics analyses. Phylogeny reconstructed using combined ND1/Cytb
sequences identified two branches in Serbian white-tailed eagles. Although the majority of
substitutions were nonsynonymous, no selective pressure was detected.
Our data suggest that combined ND1/Cytb sequence variability provides sufficient
information to be used for population comparison, population differentiation analyses
and phylogeny reconstruction, but also gives a tool to potentially identify adaptations to
different environmental conditions.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202
Determinants of CRISPR array non-canonical adaptation mechanism
Get PDFCRISPR-cas systems are incredibly diverse and currently are classified in six major types
and over 30 subtypes. Apart from their role in adaptive immunity it has been shown that
some of the CRISPR-cas subtypes are also involved in host gene regulation and even in
collateral damage leading to bacteriostatic or lethal outcomes for the host. CRISPR array
spacers direct and influence canonical and non-canonical functions of the CRISPR-cas
system together with subtype Cas proteins. Better understanding of spacer adaptation
mechanisms is crucial for uncovering intricacies of evolutionary arms race between
prokaryotes and phages.
Here we present large-scale analysis of CRISPR array spacers originating from 31845
complete bacterial genomes. All bacterial and 16388 viral genomes were retrieved using
NCBI datasets API. CRISPRidentify and CRISPRcasIdentifier tools were used for CRISPR
array, Cas genes detection and subtyping. Viral genomes were mapped to their hosts
using the latest version of the Virus-Host DB. Mapping was performed on the genus level
of the hosts phylogenetic tree. Gumbel extreme value distribution was used to determine
statistical significance of each spacer Smith-Waterman alignment score.
Differences in melting energy and GC content between identified spacers, origin bacterial
genomes and infecting bacteriophages were explored for different CRISPR-cas subtypes
and for different bacterial genera. Spacers from the extremes of the GC content distribution
were aligned to the origin bacterial and infecting phage genomes in order to determine
their origin.
GC content of the spacers was lesser than the GC content of the source bacterial genome
but greater than infecting viral genome. This observation aligns with the hypothesis that
the majority of CRISPR spacers were adapted from the bacteriophage genomes and serve
canonical function. Alignments of the spacers from GC rich distribution tail have shown
their preferential targeting of host genomes which further supports the hypothesis that
GC rich spacers originated from the bacterial genome and have non-canonical function.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202