Publications and Presentations

Pareto Diagram

QbD is known across all industries as a new management philosophy that successfully replaced the traditional business management models. The regulatory materials in the healthcare industry have been created based on the principles described in QbD programs… read more

Analytical Methodology and Toolkit for Improvement of Quality Attributes and Process Optimization of Biopharmaceuticals. Keynote talk at The 4th International Conference and Exhibition on Cell & Gene Therapy, London, UK.

The development of biologics implies thorough knowledge of protein structural and dynamic properties in order to better manage challenges related to product characterization, abnormalities — and, related to this, lack of activity, specificity, efficacy, and safety aspects… read more

Antibodies

Proteins are essential biomolecules for use in research, diagnostics, and therapy; however, toxicity and side effects, lack of activity and specificity of the manufactured proteins arise due to altered properties and factors derived from the complexity of their structures, cellular specificity of action, dynamic folding, and reactivity… read more

Crystal Structure of Human Multiple Copies in T-Cell Lymphoma-1 Oncoprotein. Protein: Structure, Function, Bioinformatics.

Human malignant T cell amplified sequence 1

Overexpression of multiple copies in T-cell lymphoma-1 (MCT-1) oncogene accompanies malignant phenotypic changes in human lymphoma cells. Specific disruption of MCT-1 results in reduced tumorigenesis, suggesting a potential for MCT-1-targeted therapeutic strategy…read more 

ML285 Affects Reactive Oxygen Species’ Inhibition of Pyruvate Kinase M2, Series: Probe Reports from the NIH Molecular Libraries Program. National Center for Biotechnology Information (U.S.A.)

The ability of all cells to regulate levels of reactive oxygen species (ROS) is vital for controlling many aspects of proliferation and survival and we have discovered that pyruvate kinase M2 (PKM2) is important for cancer cell biology. PKM2 is directly oxidized on Cys358to inhibit its catalytic activity, which allows for diversion of glucose-6-phosphate into the pentose phosphate pathway… read more

A Potent PKM2 Activator Induces Tetramerization and Reduces Tumor Formation and Size in a Mouse Xenograft Model. Series: Probe Reports from the NIH Molecular Libraries Program, National Center for Biotechnology Information.

ML265 in PKM2

Cancer cells have altered metabolic processes compared to normal differentiated cells and the expression of the M2 isozyme of pyruvate kinase (PKM2) plays an important role in this aberrant metabolism… read more

Certhrax Toxin, an Anthrax-related ADP-ribosyltransferase from Bacillus cereus. Journal of Biological Chemistry.

Certhrax Toxin in ADP-ribosyltransferase from Bacillus cereus

We identified Certhrax, the first anthrax-like mART toxin from the pathogenic G9241 strain of Bacillus cereus. Certhrax shares 31% sequence identity with anthrax lethal factor from Bacillus anthracis; however, we have shown that the toxicity of Certhrax resides in the mART domain, whereas anthrax uses a metalloprotease mechanism… read more 

Pyruvate Kinase M2 Activators Promote Tetramer Formation and Suppress Tumorigenesis. Nature Chemical Biology.

Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. The interaction of PKM2 with phosphotyrosine-containing proteins inhibits enzyme activity and increases the availability of glycolytic metabolites to support cell proliferation… read more

Crystal Structures of Human Choline Kinase Isoforms in Complex with Hemicholinium-3: Single Amino Acid Near the Active Site Influences Inhibitor Sensitivity.
Journal of Biological Chemistry.

Human choline kinase isoforms in complex with hemicholinium-3

Human choline kinase (ChoK) catalyzes the first reaction in phosphatidylcholine biosynthesis and exists as ChoKalpha (alpha1 and alpha2) and ChoKbeta isoforms. Recent studies suggest that ChoK is implicated in tumorigenesis and emerging as an attractive target for anticancer chemotherapy… read more 

High-Throughput Methods for Analyzing Transition Metals in Proteins on a Microgram Scale. Structural Proteomics, High-Throughput Methods, Series: Methods in Molecular Biology.

Transition metals are among the most common ligands that contribute to the biochemical and physiological properties of proteins. In the course of structural proteomic projects, the detection of transition metal cofactors prior to the determination of a high-resolution structure is extremely beneficial… read more

In Situ Proteolysis for Protein Crystallization and Structure Determination.
Nature Methods.

We tested the general applicability of in situ proteolysis to form protein crystals suitable for structure determination by adding a protease (chymotrypsin or trypsin) digestion step to crystallization trials of 55 bacterial and 14 human proteins that had proven recalcitrant to our best efforts at crystallization or structure determination… read more 

Escherichia coli HypA is a Zinc Metalloprotein with a Weak Affinity for Nickel. 
Journal of Bacteriology.

The hyp operon encodes accessory proteins that are required for the maturation of the [NiFe] hydrogenase enzymes and, in some organisms, for the production of urease enzymes as well. HypA or a homologous protein is required for nickel insertion into the hydrogenase precursor proteins… read more

A High-performance Liquid Chromatography Method for Determining Transition Metal Content in Proteins. Analytical Biochemistry.

HPLC metal analysis

Transition metals are common components of cellular proteins and the detailed study of metalloproteins necessitates the identification and quantification of bound metal ions. Screening for metals is also an informative step in the initial characterization of the numerous unknown and unclassified proteins now coming through the proteomic pipeline… read more

Molecular Cloning, Expression and Characterization of Three Distinctive Genes Encoding Methionine Aminopeptidase from Synechocystis sp. Strain PCC6803. Archives of Microbiology.

MetAP Metal finger

Methionine aminopeptidase, known to be encoded by single genes in prokaryotes, is a cobalt-dependent enzyme that catalyzes the removal of N-terminal methionine residues from nascent polypeptides. Three ORFs encoding putative methionine aminopeptidases from the genome of cyanobacterium Synechocystis sp. strain PCC6803 were cloned and expressed in Escherichia coliread more

Mass-Spectral Analysis of Human Interferon-gamma and Chloramphenicol Acetyltransferase I Produced in Two Escherichia coli Strains.
Journal of Biotechnology.

Mass-spectral analysis of human interferon-gamma Escherichia coli

Recombinant human interferon-gamma and chloramphenicol acetyltransferase I were isolated from two Escherichia coli strains, E. coli LE329 and E. coli XL1-blue and characterized by electrospray ionization mass spectrometry (ESI-MS). The ESI-MS analysis showed higher masses in comparison with the theoretically calculated for both proteins as well as unexpected molecular heterogeneity… read more

N-terminal Methionine in Recombinant Proteins Expressed in Two Different Escherichia coli Strains. Journal of Biotechnology.

Gamma Interferon in Ecoli strains

Two genes coding for chloramphenicol acetyltransferase and human interferon gamma, respectively, were over-expressed constitutively in two different strains of Escherichia coli (E. coli LE392 and E. coli XL1). The N-terminal amino acid analysis of the purified proteins showed that: (a) the N-terminal methionine is processed more efficiently in E. coli LE392 rather than in E. coli XL1 cells; (b) … read more

Synthesis and Antitumor Activity of Several New 4-Aroyl-1-nitrosohydrazinecarboxamides. Drugs under Experimental and Clinical Research.

4-aroyl-1-nitrosohydrazinecarboxamides

Six new analogues of nitrosoureas containing aroylhydrazine residue have been synthesized. All six compounds showed a dose-dependent in vivo activity against leukaemias L1210 and P388. Compounds I and III were soluble in water… read more

A Variant for Isolation of Serum Gangliosides. Journal of Liquid Chromatography.

The serum gangliosides are of great diagnostic importance. A step-by-step procedure for their isolation from serum is described and it includes the following stages: a) dehydration of the sample; b) total lipids extraction; c) non-polar lipids removal by preparative… read more

Differences between Some Properties of Acetylated and Nonacetylated Forms of HMG 1 Protein. Biochemical and Biophysical Research Communications.

There are data suggesting that HMG1 protein may be involved in DNA replication. Recently we have found that only the acetylated form of the protein generates tetramers, stimulates the activity of DNA polymerase alpha (EC 2.7.7.7) … read more

Chloroplast Ribosomal Protein CL 23 is Encoded in the Chloroplast Genome. FEBS Letters.

The 50 S subunit of chloroplast ribosomes was prepared from tobacco leaves. The proteins were fractionated and the N-terminal amino acid sequence of a 14 kDa protein was determined. This sequence matches the N-terminal sequence deduced from ORF55 located between ndhF and trnL on the small single-copy region of tobacco chloroplast DNA… read more

Interaction Between Glucose Diet and Ethanol on Microsomal Induction and Rat Liver Plasma Membrane Damage in Chronic Hexachlorobenzene Intoxication. 
Archives of Toxicology.

Male Wistar rats fed for 60 days a glucose diet containing 17.5 mmol hexachlorobenzene/kg show a less pronounced increase in serum parameters and microsomal cytochrome P-450 concentration and a lower decrease in liver plasma membrane 5′-nucleotidase, K+, Na+- and Mg++-adenosine triphosphatase activities than the controls fed standard diet + hexachlorobenzene. Addition of 10% ethanol to the drinking water eliminates the “glucose effect”… read more

Selection and Characterization of Amino Acid Analogues Resistant Cell Lines of Medicago Proceedings of the 2nd International Symposium of Experimental Mutagenesis in Plants.

Amino acid analogs supplied to the culture medium caused a strong inhibition of growth in a wide range of organisms. The synthesis of many amino acids in higher plants, as in bacteria, have been shown to be controlled by feedback inhibition. We synthesized Edman derivative, diphenyl-iso-thiocyanate and used to covalently modify amino acids in cellular extracts for sensitive amino acids… read more

Differences in Sheep’s Milk Composition (Whole, αs- and β- Casein) and properties due to the breed. Animal Science.

Milk degradation

Whole, αs- and β- caseins were isolated from sheep milk of three breeds grown in Bulgaria: Pleven Blackface, East Friesian, and Awassi. Water sorption andαφdpH were estimated from hysteresis curve areas obtained on acid-base titration of the studied proteins and the interbreed differences in these properties as well. The comparative profiles from the time course degradation… read more

Rat Liver Plasma Membrane Damage in Hexachlorobenzene Intoxication and its Potentiation by Ethanol. Toxicology Letters.

Chronic feeding of male Wistar rats with food containing hexachlorobenzene (HCB) at 17.5 mmol/kg induced elevation of serum amino-transferases and bilirubin content, increase of microsomal cytochrome P-450 concentration, and decrease of 5′-nucleotidase, K+, Na+- and Mg2+-adenosine triphosphatase activities in liver plasma membrane preparations… read more

The Effect of Thioacetamide on Rat Liver Plasma Membrane Enzymes and its Potentiation by Fasting. Toxicology.

The effect of thioacetamide on rat liver plasma membrane enzymes

Thioacetamide, given intraperitoneally (1.4 mmol/kg body mass) to male Wistar rats 24 h before sacrifice promoted a marked elevation of serum aminotransferases, loss of microsomal cytochrome P-450 content and a significant reduction (about 50%) of the liver plasma membrane enzymatic activities (5′-nucleotidase; K+, Na+- and Mg2+-adenosine triphosphatases; and gamma-glutamyl transferase)… read more

3D Structure Gallery

Human Non-Protein Kinases

Human Adenylate Kinase 3-Like 1, EC 2.7.4.10 (PDB: 1ZD8)
Related diseases: central nervous system disorders, ischemia,
immunological disorders.

Human Adenilate Kinase 3 Like 1

Adenylate kinase (AK), known also as phosphotransferase or myokinase,
is a key enzyme participating in cellular metabolism. AK is essential in prokaryotes, which catalyzes the interconversion of three adenine nucleotides in the cell:    MgNTP + AMP <-> MgNDP + ADP   (N = A or G)  Several AK isoforms exist in vertebrates; however, their physiological importance is not fully… read more

Nucleoside Diphosphate Kinase 3, EC 2.7.4.6 (PDB: 1ZS6)
Related diseases: cancer, anemia, rheumatoid arthritis, Alzheimer syndrome.

Protein expressed in non-metastatic cell 3 (NME3), also known as nucleoside-diphosphate kinase, nucleoside 5\’-diphosphate phosphotransferase and NM23, catalyzes the ATP gamma phosphate transfer to the NDP beta phosphate via a ping-pong mechanism, using a phosphorylated histidine intermediate in the active-site: ATP + nucleoside diphosphate <=> ADP + nucleoside… read more

Pyridoxal Kinase, EC 2.7.1.35 (PDB: 2AJP)
Related diseases: epilepsy, seizures, Parkinson syndrome, Down syndrome,
Alzheimer syndrome.

Pyridoxal kinase (PDXK, EC 2.7.1.35) catalyzes the phosphorylation of vitamin B6 (pyridoxal 5′-phosphate, PLP), a cofactor for at least 140 enzymes. PDXK is implicated in nervous system function, since many neurotransmitters such as dopamine and serotonin are synthesized by PLP-dependent enzymes.
The gene encoding three human isoforms of PDXK is located in chromosome 21,
where several complex disorders, including Down syndrome… read more

Pyruvate Kinase Isozymes M1/M2, EC 2.7.1.40 (PDB: 1ZJH, 3GQY, 3GR43H6O, 3ME3, 3U2Z) 
Related diseases: cancer, immunological disorders

Pyruvate kinase (PK, EC 2.7.1.40) catalyzes the last step of glycolysis, where the phosphoryl group of phosphoenolpyruvate (PEP) is transferred to ADP to form pyruvate and ATP, and thus participates in the primary intersections of the energy metabolism. Lately, the enzyme was linked to other diseases related to both glucose and oxygen utilization, such as diabetes, blood, and brain phenylketonuria, and angiogenesis… read more

Human Small GTPases

ADP-Ribosylation Factor 4 (PDB: 1Z6X)
Related diseases: acute erythroid leukemia, pancreatic cancer, retinal diseases.

ADP-ribosylation factor 4 (ARF4) is a member of the Ras small G protein superfamily. It is involved in membrane trafficking and mediating epidermal growth factor receptor-dependent phospholipase D2 activation. Also, ARF4 interacts with the C-terminal cytoplasmic domain of Rhodopsin… read more

ADP-Ribosylation Factor 5 (PDB: 2B6H)
Related diseases: cancer, based on mapped gene mutations.

ARF5 is a member of ADP-ribosylation factors (ARFs), ~20 kDa GTP-binding proteins. Their activity affects morphologies of organelles such as Golgi apparatus and early endosomes, and they regulate protein traffic through secretory and endocytic pathways. Biochemical studies demonstrated that one of the predominant ARF-mediated functions includes the recruitment of coat proteins to membrane surfaces, consequently facilitating vesicle budding… read more

ADP-Ribosylation Factor-like Protein 5A (PDB: 1Z6Y)
Related diseases: endocrine system diseases, Lentivirus related diseases, such as HIV.

ADP-ribosylation factor-like 5 (ARL5) belongs to the ARF family of GTP-binding proteins. The cellular function of ARL5 is currently unknown. The primary sequence of ARL5 is most similar to that of ADP-ribosylation factor-like 8 (ARL8), which is potentially involved in brain development… read more

ADP-Ribosylation Factor-like 8 (PDB: 1YZG)
Related diseases: liver transplant disorder, ischemic reperfusion injury, Whipple’s disease.

ADP-ribosylation factor (ARF) proteins are RAS-like small G proteins with molecular weights of 20 kDa. They include ARF, ARF-like (ARL) and Secretion-associated and Ras-related (SAR) proteins, all of which are collectively called the ARF family. While ARF and SAR proteins are known to function in membrane trafficking, ARL proteins are not well characterized… read more

ADP-Ribosylation Factor-like 10B (PDB: 1ZD9, 2H18
Related diseases: hyperoxia; complex V deficiency, encephalomyelitis, hepatocellular dysplasia.

ADP-ribosylation factor-like 10B (ARL10B) is a member of the ARF GTPase family currently with unknown function. The structure of a truncated mutant of ARL10B (ARL10B-N17) that lacks the N-terminal 17 amino acids has been solved at 1.7Ã resolution… read more

Related diseases: encephalomyelitis, thyroid disease, hypertrophic cardiomyopathy.

ADP-ribosylation factor-like 10C (ARL10C) is a novel small GTPase, member of the Arf family. Arfs belong to the Ras superfamily which also includes the Ras, Rho/Rac, Rab and Ran sub-families. These proteins serve as a signal transducers to regulate cellular proliferation and differentiation, actin cytoskeletal rearrangement, membrane trafficking and nuclear transport… read more 

Related diseases: Lentivirus related diseases, such as HIV.

HRB (HIV-1 Rev Binding protein), also known as hRIP (human Rev-Interacting Protein), RAB (Rev/Rex Activation domain-Binding protein), is a 60-kDa protein initially identified by yeast two-hybrid screening to interact with Rev protein of HIV-1 and assist in HIV replication. Ablation of HRB activity leads to mislocalization and aberrant accumulation of Rev-directed viral RNAs around… read more

Related diseases: cancer.

Protein exporting from the endoplasmic reticulum (ER) is the first membrane trafficking step in the secretory pathway 1. To date, all proteins may use COPII coat complexes for the initial stages of export. Sar1 belongs to the Ras-related GTPase family. It is one of the core components of COPII… read more

Related diseases: leukemia.

GTPase Activating Proteins, or the GAPs, are a family of regulatory proteins for GTPases by stimulating their GTPase (hydrolysis) activity and thus alternating signaling events. Different families of GTPases have corresponding GAPs proteins whose structure and mechanism of function varies quite differently… read more 

Related diseases: osteoporosis,  hepatitis.

GTPase Activating Proteins, or the GAPs, are regulatory proteins for Ras, Rho, Arf and Rab GTPases by stimulating the nucleotide-hydrolyzing activities of these GTPases. The Ras, Rho, Arf and Rab GTPases have their own GAPs of which the structure and reaction mechanism are different from each other… read more

Related diseases: osteoporosis,  hepatitis.
Human TBC1 domain family, member 22A

GTPase Activating Proteins, or the GAPs, are regulatory proteins for GTPases by stimulating their GTPase (hydrolysis) activity and thus alternating signalling events. Different families of GTPases have corresponding GAPs proteins whose structure and mechanism of function varies… read more

Related diseases: leukocyte adhesion deficiency, papilloma virus infections.
Human zinc finger protein 289, ID1 regulated

GTPase Activating Proteins, or the GAPs, are a family of regulatory proteins for GTPases by stimulating their GTPase (hydrolysis) activity and thus alternating signaling events. Different families of GTPases have corresponding GAPs proteins whose structure and mechanism of function varies quite differently… read more

Other Cancer Related 3D Structures

Malignant T Cell-Amplified Sequence 1 (PDB: 3R90)
Related diseases: cancer.

Genomic alteration through gene amplification has been recognized as an important event in malignant lymphomagenesis. Multiple copies in T-cell lymphoma-1 (MCT-1) is an oncogene that was originally identified in a certain lymphoma cell line, where it was found amplified… read more

Human eukaryotic translation initiation factor 4A isoform 2

The DEAD-box proteins form a widely dispersed family of proteins found in all eukaryotes and most prokaryotes. The DEAD-box proteins have been found in more than 500 proteins. The name of the family was derived from the amino-acid sequence D-E-A-D (Asp-Glu-Ala-Asp) of its Walker B motif… read more

 

Bacterial Toxins

Bacillus cereus Lethal Factor (PDB: 4FXQ)
Related diseases: foodborne illness, anthrax.

We identified Certhrax, the first anthrax-like mART toxin from the pathogenic G9241 strain of Bacillus cereusCerthrax shares 31% sequence identity with anthrax lethal factor from Bacillus anthracis; however, we have shown that the toxicity of Certhrax resides in the mART domain, whereas anthrax uses a metalloprotease mechanism. .. read more