Structural Insights into the Interaction of Cytochrome P450 3A4 with Suicide Substrates: Mibefradil, Azamulin and 6′,7′-Dihydroxybergamottin

Human cytochrome P450 3A4 (CYP3A4) is the most important drug-metabolizing enzyme. Some drugs and natural compounds can act as suicide (mechanism-based) inactivators of CYP3A4, leading to unanticipated drug-drug interactions, toxicity and therapeutic failures. Despite significant clinical and toxicological implications, the mechanism-based inactivation remains incompletely understood. This study provides the first direct insights into the interaction of CYP3A4 with three suicide substrates: mibefradil, an antihypertensive drug quickly withdrawn from the market; a semi-synthetic antibiotic azamulin; and a natural furanocoumarin, 6′,7′-dihydroxybergamottin. Novel structural findings help better understand the suicide substrate binding and inhibitory mechanism, and can be used to improve the predictability of the binding ability, metabolic sites and inhibitory/inactivation potential of newly developed drugs and other chemicals relevant to public health.

THE ROLE OF THE SITUATION IN THE STRUCTURE OF DRIVING TO SUICIDE MECHANISM

The article deals with the role of the situation in the structure of driving to suicide mechanism. The situation of driving to suicide is differentiated into stages of its development, the article also determines temporal connections of this crime and distinguishes types of driving to suicide places with the focus on the role of these elements in criminalistic characteristics of crimes as well as and their interconnections. A place of driving to suicide is a place, where preparations prior to the crime have been carried out, the crime had been committed (threats, cruel treatment, systematic humiliation of human dignity, blackmail); a place with traces and evidence of a criminal trespass (place of suicide); place of concealment of traces and evidence of the crime, instruments andfacilities of its commission, the object of a criminal trespass (staging of suicide, murder). The time of driving to suicide allows establishing the sequence and development of different processes. The situation of driving to suicide consists of non-interference and omission by individuals (relatives, colleagues, friends and others) in whose presence a criminal mistreats or humiliates the personal dignity of the victim.

Bayesian phylogeny analysis of vertebrate serpins illustrates evolutionary conservation of the intron and indels based six groups classification system from lampreys for ~500 MY

The serpin superfamily is characterized by proteins that fold into a conserved tertiary structure and exploits a sophisticated and irreversible suicide-mechanism of inhibition. Vertebrate serpins can be conveniently classified into six groups (V1-V6), based on three independent biological features - genomic organization, diagnostic amino acid sites and rare indels. However, this classification system was based on the limited number of mammalian genomes available. In this study, several non-mammalian genomes are used to validate this classification system, using the powerful Bayesian phylogenetic method. This method supports the intron and indel based vertebrate classification and proves that serpins have been maintained from lampreys to humans for about 500 MY. Lampreys have less than 10 serpins, which expanded into 36 serpins in humans. The two expanding groups V1 and V2 have SERPINB1/SERPINB6 and SERPINA8/SERPIND1 as the ancestral serpins, respectively. Large clusters of serpins are formed by local duplications of these serpins in tetrapod genomes. Interestingly, the ancestral HCII/SERPIND1 locus (nested within PIK4CA) possesses group V4 serpin (A2APL1, homolog of α2-AP/SERPINF2 ) of lampreys; hence, pointing to the fact that group V4 might have originated from group V2. Additionally in this study, the phylogenetic history and genomic characteristics of vertebrate serpins were revisited.

Bayesian phylogeny analysis of vertebrate serpins illustrates evolutionary conservation of the intron and indels based six groups classification system from lampreys for ~500 MY

The serpin superfamily is characterized by proteins that fold into a conserved tertiary structure and exploits a sophisticated and irreversible suicide-mechanism of inhibition. Vertebrate serpins can be conveniently classified into six groups (V1-V6), based on three independent biological features - genomic organization, diagnostic amino acid sites and rare indels. However, this classification system was based on the limited number of mammalian genomes available. In this study, several non-mammalian genomes are used to validate this classification system, using the powerful Bayesian phylogenetic method. This method supports the intron and indel based vertebrate classification and proves that serpins have been maintained from lampreys to humans for about 500 MY. Lampreys have less than 10 serpins, which expanded into 36 serpins in humans. The two expanding groups V1 and V2 have SERPINB1/SERPINB6 and SERPINA8/SERPIND1 as the ancestral serpins, respectively. Large clusters of serpins are formed by local duplications of these serpins in tetrapod genomes. Interestingly, the ancestral HCII/SERPIND1 locus (nested within PIK4CA) possesses group V4 serpin (A2APL1, homolog of α2-AP/SERPINF2 ) of lampreys; hence, pointing to the fact that group V4 might have originated from group V2. Additionally in this study, the phylogenetic history and genomic characteristics of vertebrate serpins were revisited.

Anticoagulative and anticomplementary activity of endogenous inhibitor preparation from hepatopancreas of red king crab (paralithosed camtschaticus) towards human blood

Serpins (SERine Protease INhibitors) - is large and diverse group of proteins with similar structures, which can inhibit both serine and cysteine proteases by an irreversible suicide mechanism. A novel serpin from hepatopancreas of Red King Crab (Paralithosed camtschaticus) was obtained and was studied its effect on the process of human blood plasma clotting. The investigated serpin shows a noticeable anticoagulative activity, which increases dramatically in the combined action with heparine. Though the inhibitor has almost no effect on thrombin, it inhibits C1s (C1-esterase). We studied the action of the serpin from P. camtschaticus on C1s via its competitive inhibition by C1 inhibitor and the novel enzyme. The calculated inhibition constant of the serpin from P. camtschaticus towards C1s is 2,02±0,71 М. Unlike C1 inhibitor, the novel serpin from P. camtschaticus doesn`t suppress fibrinolysis and at the same time prevents blood clotting. These features may be of interest for medical purposes.

Are caspases involved in the death of cells with a transcriptionally inactive nucleus? Sperm and chicken erythrocytes

We show that mouse sperm die spontaneously within 1–2 days in culture and that treatment with either staurosporine (STS) and cycloheximide (CHX) or a peptide caspase inhibitor does not accelerate or delay the cell death. Chicken erythrocytes, by contrast, are induced to die by either serum deprivation or treatment with STS and CHX, and embryonic erythrocytes are more sensitive than adult erythrocytes to both treatments. Although these erythrocyte deaths display a number of features that are characteristic of apoptosis, they are not blocked, or even delayed, by peptide caspase inhibitors, and most of the cells die without apparently activating caspases. A small proportion of the dying erythrocytes do activate caspase-3, but even these cells, which seem to be the least mature erythrocytes, die just as quickly in the presence of caspase inhibitors. Our findings raise the possibility that both mouse sperm and chicken erythrocytes have a death programme that may not depend on caspases and that chicken erythrocytes lose caspases as they mature. Chicken erythrocytes may provide a useful ‘stripped down’ cell system to try to identify the protein components of such a death programme, which may serve to back-up the conventional caspase-dependent suicide mechanism in many cell types.