High Degree Of Affinity Research Articles

Simulation study of jet trajectory based on Runge-Kutta method

Abstract To solve the problem of not being able to accurately spray to the designated target point due to wind and distance factors when firefighting at a long distance, this paper adopts the Runge-Kutta algorithm to simulate the trajectory of the fire cannon, and establishes the mathematical model of the trajectory of the jet under the ideal state and the trajectory of the jet under the influence of the wind. We carry out three-dimensional visualization research and compare the results of the simulation experiment with the real experimental data, and the range, height, and trajectory have a high degree of fit. Comparing the simulation results with the real experimental data, the range and height of the jet and the trajectory have a high degree of affinity, and adding the wind power value panel in the visualization interface can simulate the changes of the jet trajectory under different wind speeds and wind power and effectively simulate the trajectory of the jet under natural conditions.

Molecular epidemiology and resistance mechanisms of tigecycline-non-susceptible Acinetobacter baumannii isolated from a tertiary care hospital in Chongqing, China.

We studied 34 isolates of Tigecycline-Non-Susceptible A. baumannii (TNAB) obtained from clinical specimens at a large tertiary care hospital in Chongqing, China. These 34 strains belonged to 8 different clones including ST195 (35.3%) and ST208 (17.7%). EBURST analysis found that these 8 ST types belonged to the Clonal Complex 92. Tigecycline resistance-associated genes adeR, adeS, adeL, adeN, rrf, rpsJ, and trm were detected in most strains. The expression level of the resistance-nodulation-cell division (RND) efflux pumps in TNAB strains was higher than the reference strain ATCC19606. 58.8% of strains had a decrease in the tigecycline minimum inhibitory concentration (MIC) after the addition of carbonyl cyanide 3-chlorophenylhydrazone (CCCP). The TNAB strains in our hospital have a high degree of affinity and antibiotic resistance. Regular surveillance should be conducted to prevent outbreaks of TNAB epidemics.

A61 TARGETING A GUANINE QUADRUPLEX IN THE HEPATITIS B VIRUS COVALENTLY CLOSED CIRCULAR DNA GENOME USING SINGLE DOMAIN ANTIBODIES

Abstract Background The establishment of chronic HBV infection in over 297 million people is due in part through the virus’ highly stable covalently closed circular DNA (cccDNA) persisting in the nuclei of infected hepatocytes. Thus, curing HBV will require direct targeting of the minigenome and detailed structural knowledge of the cccDNA to determine vulnerabilities. Purpose Previous studies have demonstrated that human protein Sp1 interacts with the HBV genome at the preCore/Core promoter - a critical interaction for viral replication. We have recently discovered that the Sp1-binding region of HBV pre-Core forms a highly ordered G-quadruplex (G4) secondary structure which presents a novel therapeutic anti-HBV target. Method Using phage display technologies, we have identified 11 G-quadruplex binding single domain antibodies that can target the G4 present within the cccDNA. Using recombinant protein expression we characterized the strongest binder (S10) and its interaction with a 22nt HBV pre-Core G-quadruplex forming oligo. Using MicroScale Thermophoresis (MST), the binding affinity (KD) between S10 and the target G4 was determined to be ~218 nM, which is 100x stronger for folded G4 versus unfolded oligos of the same sequence. Using oligos of (~60-90%) sequence similarity, it was observed that S10 has a KD for the target G4 that was at least 10x higher than similar sequences. To determine the effect of S10 on HBV replication, we have transduced the HepG2-NTCP-A3 cell line to express these sdAbs and are in the process of evaluating antiviral effects and target specificity for cccDNA. Result(s) Using biophysical in-vitro approaches, S10 has shown a great potential in being able to discriminate against different G4’s, while having a high degree of affinity as well as high complex stability. Conclusion(s) The ability of these G4 binding single domain antibodies to discriminate between different sequences or secondary DNA structure provides an insight into how they can be exploited in future HBV therapeutic strategies. Disclosure of Interest None Declared

A Novel ssDNA Aptamer Targeting Carcinoembryonic Antigen: Selection and Characterization.

Simple SummaryCarcinoembryonic antigen (CEA), a well-known cancer biomarker that is used to diagnose various cancers, notably colorectal cancer, was used as a target to select and characterize single-stranded DNA aptamers. These short and compact nucleic acid sequences, known as aptamers, are becoming ever more popular in both therapeutics and diagnostics. In this study, SELEX and NGS techniques were employed in the aptamer selection processes. Aptamer characterization techniques, such as enzyme-linked oligonucleotide assay (ELONA), dot blot, and bioinformatics assays, were conducted to observe the affinity binding of the selected aptamer to the target protein. Confocal microscopy results demonstrated how the fluorescently labelled aptamer sequence recognized CEA, which is expressed in HT-29 human colon adenocarcinoma cell line. These findings show that the selected aptamer has the potential to be employed as a recognition component in rapid detection systems, such as aptasensors.One of the major causes of a drastically shorter life expectancy and one of the most prevalent diseases in the world today is cancer. Given the data on the rise in cancer cases throughout the world, it is obvious that, despite the diagnostic techniques currently being used, there is a pressing need to develop precise and sensitive techniques for early diagnosis of the disease. A high degree of affinity and specificity towards particular targets is maintained by the short nucleic acid molecules known as aptamers. Aptamers outperform antibodies due to their unique benefits, such as their simplicity in synthesis and modification, lack of toxicity, and long-term stability. Utilizing an accurate recognition element and a robust signal transduction mechanism, molecular diagnostics can be extremely sensitive and specific. In this study, development of new single-stranded DNA aptamers against CEA for use in cancer diagnostics was accomplished using SELEX and NGS methods. As a result of 12 iterative SELEX rounds, nine aptamer candidates against CEA were developed. NGS comparative analysis revealed that round twelve had an enriched number of aptamers that were specifically bound, as opposed to round eight. Among the selected nine sequences characterized by bioinformatics analysis and ELONA, an aptamer sequence with the highest specificity and affinity for the target protein was identified and further examined. Aptamer sequence (6) was screened in a concentration-dependent assay, specificity analysis was performed, and its potential secondary and tertiary structures were predicted, which enabled us to test one of the possible putative interactions with CEA. Finally, aptamer sequence (6) labelled with a Cy5 fluorescent tag was used in confocal microscopy to observe its binding towards the CEA expressed in HT-29 human colon adenocarcinoma cell line.

Mechanisms of action of cytoplasmic microRNAs. Part 3. TNRC6-associated mechanism of miRNA-mediated mRNA degradation

The scientific review presents the mechanisms of action of cytoplasmic miRNAs, namely posttranscriptional silencing: the TNRC6-associated mechanism of miRNA-mediated mRNA degradation. To write the article, information was searched using databases Scopus, Web of Science, MedLine, PubMed, Google Scholar, EMBASE, Global Health, The Cochrane Library, CyberLeninka. It is known that in the cytoplasm of cells in cases of short region, miRNA complementarities cause posttranscriptional silencing, using the first of the main molecular mechanisms: the TNRC6-associated mechanism of miRNA-mediated mRNA degradation. Mammalian AGO proteins have been shown to contain the conserved m7G-cap-binding protein motif (known as the MID domain), which is required to induce microRNA-mediated translation repression. After binding of this AGO motif to microRNAs, TNRC6 proteins (GW182) are recruited that, in turn, recruits various proteins (PABPC1, PAN3 and NOT1) involved in the induction of the target gene silencing. The authors state that tryptophan residues, which are placed in the hydrophobic pockets of TNRC6 protein partners, cause a high degree of affinity and specificity of interactions. Scientists believe that the TNRC6 protein when interacting with AGO proteins can simultaneously use three GW/WG repeats (motif 1, motif 2 and hook motif), which are located in the Argonaute-binding domain. Therefore, the TNRC6 protein can bind to three AGO molecules simultaneously. TNRC6 proteins are known to be PABP-interacting proteins whose interaction with PABP is mediated by conservative PABP-binding motif 2. TNRC6 proteins have been shown to interact with the cytoplasmic PABPC1 protein during mRNA translation and stabilization. It is shown that the CCR4-NOT protein complex is a highly conserved multifunctional multiprotein formation having 3’-5’-exoribonuclease activity, due to which it controls mRNA metabolism. Thus, the TNRC6-associated mechanism of miRNA-mediated mRNA degradation in the cytoplasm of the cell causes posttranscriptional silencing. In this mechanism, there is an interaction of TNRC6 with PABPC1 protein, recruitment of deadenylating complexes PAN2-PAN3 and CCR4-NOT by the TNRC6 proteins.

Ovarian cancer and KiSS-1 gene expression: A consideration of the use of Kisspeptin plus Kisspeptin aptamers in diagnostics and therapy

Gynaecological cancers continue to present a significant health burden upon the health of the global female population. This deficit is most prominent with ovarian cancer which possesses the lowest survival rate compared to all other cancers occurring within this anatomical region, with an annual UK-mortality of 7,300. The poor tolerability and selectively of the treatment options that are currently available is likely to have contributed to this high mortality rate thus, demonstrating the need for the development of enhanced therapeutic approaches. Aptamer technology would involve the engineering of specifically sequenced oligonucleotide chains, which bind to macromolecular targets with a high degree of affinity and selectively. Recent in-vitro studies conducted upon the clinical utility of this technique have supported its superiority in targeting individual therapeutic drug targets compared to various other targeting moieties currently within therapeutic use such as, monoclonal antibodies. For this reason, the employment of this technique is likely to be favourable in reducing the incidence of non-specific, chemotherapy-associated adverse effects. Kisspeptin is a naturally expressed polypeptide with an established role in the development of the reproductive system and other proposed roles in influencing the ability of ovarian cancer growths to exhibit the metastasis hallmark. This distinctive feature would indicate the potential for the manipulation of this pathway through the application of aptamer structures in developing a novel prophylactic strategy and improve the long-term outcome for ovarian cancer patients.

An approach for prediction of flood hydrograph at outlet of an ungauged basin using modified dynamic wave model

ABSTRACT Floods are the most disastrous natural hazards, but the prediction of flood hydrographs is hindered due to the lack of Hydro-meteorological data. This paper presents an approach for flood routing of the ungauged basin using rainfall-runoff and Dynamic wave models. Specifically, the SCS-CN rainfall-runoff model is employed to obtain the inflow and lateral inflow hydrographs of the ungauged sub-basins, and a Modified Dynamic Wave Model is employed to anticipate the flood hydrograph at the outlet of the ungauged basins. The Modified Dynamic Wave Model for the parabolic channel (MDWMP) has been numerically solved using an implicit finite difference scheme considering both continuity and momentum equations. The method is tested in hypothetically considered ungauged Kulsi River Basin, a mountainous basin in Northeastern India, and the results obtained are compared with the observed data at the outlet of the Kulsi Basin. The approach is validated by considering four parameters RMSE, Peak flow Error, Peak time Error, and Total Volume Error. A high degree of affinity (i.e. peak flow error below 6%) was observed between the predicted outflow hydrograph and the observed hydrograph, which verified the satisfactory applicability of the developed approach to anticipate the flood hydrograph of an ungauged basin.

Role of Ion Chemistry and Hydro-Geochemical Processes in Aquifer Salinization—A Case Study from a Semi-Arid Region of Haryana, India

In the present study, a total of sixty groundwater samples, twenty each for the pre-monsoon, monsoon and post monsoon seasons of 2018, were collected from selected locations in the Mewat district of Haryana, India. Electrical conductivity (EC) was measured at the site and total dissolved solids (TDS) were estimated. Samples were analysed for anions (chloride, sulphate, and bicarbonate) and cations (calcium, potassium, magnesium, and sodium). Multiple regression analysis was performed to analyse the data and report the dominant ions. Piper trilinear diagram and Gibbs plots were used to find out the water type and the factors controlling the chemistry of the groundwater, respectively. The saturation index of CaCO3, CaSO4 and NaCl was determined, using the PHREEQC MODEL. Sodium and calcium among cations, and chloride among the anions, had the highest degree of affinity and strong significance for all three seasons. The calcium–chloride water type dominated for all three seasons and Gibbs plot depicted that most of the Na+/Na+ + Ca2+ and Cl−/Cl− + HCO3− ratios show the weathering of rocks to form minerals as the major reason behind the ionic chemistry of the groundwater. The highest level of dissolution is encountered in the case of NaCl, followed by CaSO4, whereas CaCO3 depicts precipitation. The geochemical aspects of weathering, evaporation and ion exchange are the major processes responsible for high salinity, and anthropogenic activities are leading to its expansion. The findings from this study will be useful in management and remediation of groundwater salinity of the region.

Degree of affinity among dairy heifers affects access to feed supplementation

The aim of this study was to evaluate the influence of social position and degree of affinity among heifers attempting to access limited feed supplement. This study was divided into two stages: (1) observation of behaviour at the paddock and (2) arena tests. In stage 1, we measured socio-positive interactions (licking and frequency of proximity among animals) and agonistic interactions in a herd of nineteen dairy heifers. The frequency of proximity between animals of different dominance ranks was used as a measure of the degree of affinity. This measure was considered for two arena tests. In the first we compared pairs with high (PHda, n = 7) and low (PLda, n = 7) degree of affinity. In the second arena test a third neutral middle-ranking animal was included, and we compared triads with a pair with high (THda, n = 7) or low (TLda, n = 7) degree of affinity. In the arena tests, feed supplementation was offered to pairs and triads, and latency to access food, time spent at the feeder, and agonistic interactions were measured. The degree of affinity influenced (p < 0.05) the time at the feeder of the low-ranking animals at the arena test with pairs, but not with triads. Low-ranking animals remained longer (10.36 vs. 8.96 min; p ≤ 0.02) at the feeder in pairs with high degree of affinity. However, latency wasn’t affected by rank nor by affinity among pairs (p ≥ 0.13) High-ranking animals remained longer at the feeder in both treatments (p ≤ 0.02). The latency of low-ranking animals to access the food tended (p = 0.084) to be lower in triads of low degree of affinity. Despite degree of affinity, low-ranking animals were the main victims of aggressive interactions throughout the experimental period. We concluded that the degree of affinity among animals could influence access to food, but that it could not overcome the effects of social dominance.

Developing High Sensitivity/Specificity Detection Systems for Studying Protein Interactions

The study of proteomics succeeds the deciphering of the genetic code; this growing burgeoning area is set to dominate scientific research well into the next decade. Current tools employed in studying protein-protein interactions include antibodies, non-protein scaffolds, fluorescence imaging, split enzymes and the relatively new tool termed Adhirons designed by researchers at Leeds University. Antibodies have been used extensively in protein studies due to the high degree of affinity and specificity however the rise in cost and the length of time required to make antibodies has fuelled efforts to find better alternatives. In this work we report whether Adhirons owing to their small size and high stability can be adapted to assay interactions in cells. It will explore whether current tools widely used in protein studies can debunk the davinchi code for protein-protein interactions. Most biological processes are governed by protein interactions and at the heart of most disease states particularly cancer lies a signalling cascade triggered by a plethora of protein interactions. We review current research into proteomics to evaluate and appreciate the work achieved thus far by international scientist crossing east and west divide. The journey into proteomics has already begun and at the present juncture has made significant milestones.

Epitope Mapping Using Yeast Display and Next Generation Sequencing.

Monoclonal antibodies are the largest class of therapeutic proteins due in part to their ability to bind an antigen with a high degree of affinity and specificity. A precise determination of their epitope is important for gaining insights into their therapeutic mechanism of action and to help differentiate antibodies that bind the same antigen. Here, we describe a method to precisely and efficiently map the epitopes of multiple antibodies in parallel over the course of just several weeks. This approach is based on a combination of rational library design, yeast surface display, and next generation DNA sequencing and provides quantitative insights into the epitope residues most critical for the antibody-antigen interaction. As an example, we will use this method to map the epitopes of several antibodies that neutralize alpha toxin from Staphylococcus aureus.

Personnel Selection and Person-Organization, Job-Fit, Implication for Employees’ Performance in Nigeria

Person- organization-job-fit can be seen as the degree of congruence between a person and the organization, on the one hand, and between a person and his job, on the other. This in turn determines employee performance. Studies of personnel selection process generally agree that how personnel are selected into organization has a high degree of affinity with person-organization-job fit. The present study has extensively tried to establish the compatibility between a person and the organization basing on the extent to which a person and the organization share similar characteristics and/or meet such other needs. The study has also tried to establish a relationship between the process of selecting an employee and the organization-person fit. After extensive review of literature, the paper concludes that the selection/recruitment process to a large extent determines organization-person fit, on the one hand, and person-job fit, on the other.

Naphthalene diimide-polyamine hybrids as antiproliferative agents: Focus on the architecture of the polyamine chains

Naphthalene diimides (NDIs) have been widely used as scaffold to design DNA-directed agents able to target peculiar DNA secondary arrangements endowed with relevant biochemical roles. Recently, we have reported disubstituted linear- and macrocyclic-NDIs that bind telomeric and non-telomeric G-quadruplex with high degree of affinity and selectivity. Herein, the synthesis, biological evaluation and molecular modelling studies of a series of asymmetrically substituted NDIs are reported. Among these, compound 9 emerges as the most interesting of the series being able to bind telomeric G-quadruplex (ΔTm = 29 °C at 2.5 μM), to inhibit the activity of DNA processing enzymes, such as topoisomerase II and TAQ-polymerase, and to exert antiproliferative effects in the NCI panel of cancer cell lines with GI50 values in the micro-to nanomolar concentration range (i.e. SR cell line, GI50 = 76 nM). Molecular mechanisms of cell death have been investigated and molecular modelling studies have been performed in order to shed light on the antiproliferative and DNA-recognition processes.

Precise and Efficient Antibody Epitope Determination through Library Design, Yeast Display and Next-Generation Sequencing

The ability of antibodies to bind an antigen with a high degree of affinity and specificity has led them to become the largest and fastest growing class of therapeutic proteins. Clearly identifying the epitope at which they bind their cognate antigen provides insight into their mechanism of action and helps differentiate antibodies that bind the same antigen. Here, we describe a method to precisely and efficiently map the epitopes of a panel of antibodies in parallel over the course of several weeks. This method relies on the combination of rational library design, quantitative yeast surface display and next-generation DNA sequencing and was demonstrated by mapping the epitopes of several antibodies that neutralize alpha toxin from Staphylococcus aureus. The accuracy of this method was confirmed by comparing the results to the co-crystal structure of one antibody and alpha toxin and was further refined by the inclusion of a lower-affinity variant of the antibody. In addition, this method produced quantitative insight into the epitope residues most critical for the antibody–antigen interaction and enabled the relative affinities of each antibody toward alpha toxin variants to be estimated. This affinity estimate serves as a predictor of neutralizing antibody potency and was used to anticipate the ability of each antibody to effectively bind and neutralize naturally occurring alpha toxin variants secreted by strains of S. aureus, including clinically relevant strains. Ultimately this type information can be used to help select the best clinical candidate among a set of antibodies against a given antigen.

90 Intra-accumbens injection of a dopamine aptamer abates MK-801-induced cognitive dysfunction in a model of Schizophrenia

Aptamers are short, single-stranded sequences of DNA or RNA, which exhibit conformationally unique three-dimensional structures. Aptamer binding to target molecular substrates (from small molecules to proteins) demonstrates a high degree of affinity and specificity, in an analogous fashion to antigen–antibody binding. Aptamers offer several advantages over antibodies, including highly reproducible synthesis and ease of modification. Though the characteristics of aptamers and their applicability within the central nervous system show great potential, research in this area is limited. In the present work, the ability of a dopamine-binding DNA aptamer to regulate MK-801-induced cognitive deficits when injected into the nucleus accumbens was investigated. Systemic administration of the non-competitive NMDA-receptor antagonist, MK-801, has been proposed to model cognitive deficits similar to those seen in patients with schizophrenia. Rats were trained to bar press for chocolate pellet rewards then randomly assigned to receive an intra-accumbens injection of a DNA aptamer (200 nM; n = 7), tris buffer (n = 6) or a randomized DNA oligonucleotide (n = 7). Animals were then treated systemically with MK-801 (0.1 mg/kg) and tested for their ability to extinguish their bar-pressing response. Injection of 200 nM dose of the dopamine aptamer reversed this MK-801-induced elevation in lever pressing to levels as seen in rats not treated with MK-801. Tests for activity showed that the aptamer did not impair locomotor activity. Results demonstrate the in vivo utility of DNA aptamers as tools to investigate neurobiological processes in preclinical animal models of mental health disease.

Derivation of Flood Hydrographs for Ungauged Upstream Subwatersheds Using a Main Outlet Hydrograph

Understanding watershed systems is a necessary prerequisite for comprehensive management of the overall basin. Applying simple hydrologic techniques, for which fundamental information is available, is a feasible solution for determination of subwatershed contributions to the generation of total runoff. In this study, an attempt was made to develop an analytical procedure through which the outflow hydrographs could be determined for each subwatershed/area. The technique was based on reverse application of the Muskingum hydrologic routing model and the Area-CN role factor. The technique introduced was then tested to determine if it could solve flood routing problems associated with data for which the input hydrographs were known. A high degree of affinity (i.e., relative error below 32%) was observed between the estimated hydrographs and the recorded data, which verified the satisfactory applicability of the developed approach for estimation of the runoff generated by each subwatershed/area.

Polyhydroxyalkanoate chip for the specific immobilization of recombinant proteins and its applications in immunodiagnostics.

In this study, a novel strategy was developed for the highly selective immobilization of proteins, using the polyhydroxyalkanoate (PHA) depolymerase substrate binding domain (SBD) as an active binding domain. In order to determine the appropriacy of this method for immunodiagnostic assays, the single-chain antibody (ScFv) against the hepatitis B virus (HBV) preS2 surface protein and the severe acute respiratory syndrome coronavirus (SARS-CoV) envelope protein (SCVe) were fused to the SBD, then directly immobilized on PHA-coated slides via microspotting. The fluorescence-labeled HBV antigen and the antibody against SCVe were then utilized to examine specific interactions on the PHA-coated surfaces. Fluorescence signals were detected only at the spotted positions, thereby indicating a high degree of affinity and selectivity for their corresponding antigens/antibodies. Furthermore, we detected small amounts of ScFv-SBD (2.7 ng/mL) and SCVe-SBD fusion proteins (0.6 ng/mL). Therefore, this microarray platform technology, using PHA and SBD, appears generally appropriate for immunodiagnosis, with no special requirements with regard to synthetic or chemical modification of the biomolecules or the solid surface.

In vitro formation of holes on the inner perivitelline layer of quail ovum by chicken spermatozoa

ABSTRACTThe aim of the present study was to examine whether chicken semen can be substituted for quail semen to conduct in vitro experiments on fertilization. Chicken spermatozoa was incubated with the inner perivitelline layer (IPL) isolated from the largest follicle in the quail ovary under in vitro conditions. The perforation of chicken and quail spermatozoa were assessed by counting the number of all visible holes in the pieces of IPL. No difference was found in the number of holes formed by the chicken sperm and quail IPL interaction compared with that between intraspecies. In addition, the number of holes in the IPL was significantly increased with the increase in sperm concentration from 1 × 106 to 8 × 106 sperm/mL (P &lt; 0.05). Interestingly, after treatment of chicken spermatozoa with 2.5 mg/mL of the solubilized quail IPL followed by incubation with the intact chicken IPL, the number of holes in the intact chicken IPL was significantly decreased as compared with that of spermatozoa treated without the solubilized IPL (P &lt; 0.05). This indicates that sperm receptors in the solubilized quail IPL and binding ligands on the chicken spermatozoa would find each other and bind, forming complexes and these complexes blocked the interaction between chicken spermatozoa and intact chicken IPL. These results show that: (i) chicken spermatozoa possess the penetrability into quail IPL; and (ii) a high degree of affinity via the receptor interactions exists between chicken spermatozoa and quail IPL. Therefore, it appears that the substitution of chicken semen for quail semen is possible to use as an in vitro technique to examine the sperm‐IPL interaction during fertilization in quail.

Kinetic study of monophenol and o-diphenol binding to oxytyrosinase

The complex reaction mechanism of tyrosinase involves three enzymatic forms, two overlapping catalytic cycles and a dead-end complex. The deoxytyrosinase form binds oxygen with a high degree of affinity, K s O 2 = 46.6 ± 2.4 μM. The mettyrosinase and oxytyrosinase forms bind monophenols and o-diphenols, although the former is inactive on monophenols. Analytical expressions for the catalytic and Michaelis constants of tyrosinase towards phenols and o-diphenols have been derived. Thus, the Michaelis constant of tyrosinase towards monophenols ( K m M ) and o-diphenols ( K m D ) are related with the catalytic constants for monophenols ( k cat M ) and o-diphenols ( k cat D ) , respectively, and with the binding rate constants of the oxytyrosinase form with these substrates ( k +4 and k +6, respectively), by means of the expressions K m M = k cat M / k + 4 and K m D = k cat D / k + 6 . From these expressions, we calculate the values of the binding rate constant of oxytyrosinase to the substrates (monophenols and o-diphenols) for tyrosinases from different biological sources, and reveal that the o-diphenols bind more rapidly to oxytyrosinase than the monophenols. In addition, a new kinetic constant K m D ( M ) = k cat M / 2 k 6 (the Michaelis constant for o-diphenol in the monophenolase activity), is derived and determined. Thus, it has been shown that tyrosinase has apparently higher affinity towards o-diphenols in its monophenolase than in its diphenolase activity.

Cysteine proteinases and cystatins

This review describeds the definition, localization, functions and examples of cysteine proteinases and their protein inhibitors in vertebrate, non-vertebrate animals and plants. These inhibitors are related with defense mechanisms of plant against pests. It also describes the factors involved in the specific cysteine proteinase-cystatin interaction and high degree of affinity and large specificity in this interaction which are not only represented by the compatibility between amino acid residues of the active site involved in catalysis, but also of all amino acid residues that participante in the enzyme-inhibitor interaction.