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INTRODUCTION
Published in David M. Gibson, Robert A. Harris, Metabolic Regulation in Mammals, 2001
David M. Gibson, Robert A. Harris
bach enzyme catalyst possesses a region designated the "active site" where the specific rcactants (substrates) bind tightly but revcrsibly. Depending on the concentrations of the substrates and the enzyme in solution the amount of the enzyme-substrate complcx formed in this very rapid equilibration will usually determine the rate at which the reaction will proceed. The active sites are physical templates constructed with certain of the projecting amino acid side chains of the enzyme (figure 1.4). Variously chargée! or hydrophobic in nature, the three-dimensional arrangements of side chains not only restrict what particular substrates can bind but also define the precise orientations of the bound substrates to each other. Indeed the catalytic efficiency of an enzyme depends on the exquisite alignment of the interacting domains of the substrates. The coupling of one enzyme system with another through a common intermediate also depends on stereospecilic binding of substrates (and products) to cognate enzymes. litis is facilitated if sequential enzymes are placed near each other, or bound to each other. The extreme is DNA/RNA template-ordered synthetic steps (Figures 1.5 and 1.6).
Future Strategies for Commercial Biocatalysis
Published in Peter Grunwald, Pharmaceutical Biocatalysis, 2019
Robert E. Speight, Karen T. Robins
In order to assemble one-pot cascades such as the example in Fig. 1.2, as with any biocatalytic reaction, it is necessary to choose the most appropriate enzymes to start with based on clear selection criteria as well as compatibility with other enzymes in the cascade. Selection criteria are often focused on enzyme kinetics and catalytic efficiency. Other key criteria are enzyme stability and optima of activity under the reaction conditions (e.g., temperature, pH, and salt concentrations), substrate or product inhibition (including products downstream in the cascade), cofactor requirements and options for cofactor recycling either by additional enzymes or by enzymes within the cascade. An example of this cofactor recycling within a cellular pathway or cascade system is the cycling of NAD+ and NADH in the production of ethanol by Saccharomyces cerevisiae through the Embden–Meyerhof pathway. In this system the transformation of glyceraldehyde-3-phosphate to 1,3-diphosphoglycerate converts NAD+ to NADH whereas the final step to ethanol from acetaldehyde catalysed by alcohol dehydrogenase converts NADH to NAD+, meaning that the system is self-sufficient for this cofactor. In the sugar synthesis example described above, ATP and NAD+ recycling was achieved through two additional enzymes that were not part of the pathway or cascade but were present solely for recycling. A further key criterion is the commercial availability of an enzyme or the ability to readily produce the enzyme in an active and soluble form in a recombinant microbial production system.
Modulation of Tumor Matrix by Components of the Plasminogen-Plasmin System
Published in Róza Ádány, Tumor Matrix Biology, 2017
Following scu-PA binding to the receptor, its conversion to the two-chained form by Pn is greatly facilitated, estimated to be 50-fold greater than the in vivo action of Pn.90 The receptor-bound tcu-PA also converts Pg to Pn at a greater rate.90 The second-order catalytic efficiency of u-PA in Pg activation is increased sixfold on the cell surface due to a decrease of the Km from 25 μM in the fluid phase to 0.7 μM in the presence of cells.90,92 The inhibition by both PAI-1 and PAI-2 is decreased by about 40% after u-PA is bound to its receptor.90–92 Mapping studies on cells showed that the u-PA, bound to its receptor, is localized at focal contacts of cells, where they facilitate proteolysis at the adhesion sites during cell movement.95,96 Further analysis revealed that phosphorylation of the u-PA occurs during transportation to the cell surface from the cytosol.97,98 The six tyrosine residues in the ATF are phosphorylated by the src pp60 endogenous protein kinase and serine residues by protein kinase C, respectively. Whether this focal contact with the cytoplasm may initiate any signal transduction is the subject of intense investigation at the time of this review. The phosphorylated u-PA has different enzymatic properties from the non-phosphorylated form. Its catalytic effect of Pg expressed as Kcat, was fivefold that of the nonphosphorylated form, but the Km was 70-fold, resulting in a tenfold lower catalytic efficiency. However, the inhibition by PAI-1 and PAI-2 was much less in the phosphorylated form, approximately 35-fold less than inhibition of the non-phosphorylated u-PA.
A review on the mechanistic details of OXA enzymes of ESKAPE pathogens
Published in Pathogens and Global Health, 2023
Fatma Gizem Avci, Ilgaz Tastekil, Amit Jaisi, Pemra Ozbek Sarica, Berna Sariyar Akbulut
Substrate profiles of OXA-10, OXA-23, and OXA-48 can be determined by measuring the kinetic parameters such as kcat and Km. As a general rule, the kcat/Km ratio provides an idea about the catalytic efficiency of the enzymes, e.g. a high kcat/Km value would indicate high catalytic efficiency despite low kcat and Km values. Thus, the actual catalytic efficiency of a β-lactamase against a β-lactam antibiotic is commonly determined by evaluating kcat and kcat/Km values together [80]. Following this, the kinetic values presented in Tables 4, 5, and 6 clearly show that OXA-10, OXA-23, and OXA-48 display diverse substrate specificities. However, overall, they hydrolyze penicillins more efficiently when compared to other classes of β-lactams. Despite their higher affinity for carbapenems (in the nanomolar range for selected antibiotics), their hydrolysis is very slow. This demonstrates that hydrolysis of cephalosporins and monobactams is not as efficient as penicillins.
Nanoparticles in nanomedicine: a comprehensive updated review on current status, challenges and emerging opportunities
Published in Journal of Microencapsulation, 2021
Heidi Mohamed Abdel-Mageed, Nermeen Zakaria AbuelEzz, Rasha Ali Radwan, Saleh Ahmed Mohamed
Intriguingly, nanoparticles with ‘enzyme-mimetic’ activity have been studied as alternatives to natural enzymes. Catalytically active nanomaterials specifically in Nanohybrid iron oxide NP formulation and preparation allowed the introduction of enzyme mimetics, possessing peroxidase, oxidase, superoxide dismutase and catalase-like activities (nanozymes) (Singh 2019). The field of nanozymes offers promising new biomedical applications, from biofilm disruption to neurodegeneration protection and cancer prevention however, it is still in its infancy (Cormode et al.2018). Studies published are remarkable nonetheless several questions still are unanswered, which endorses further research pursue. The interwoven relationship between catalytic efficiency, therapeutic activity and biocomptability is yet to be resolved. High-performance nanozymes and highly selective nanozymes are to be developed to match catalytic efficiency natural enzymes. Also problems with batch-to-batch variation in size and shape of nanoparticles/nanozymes, and thus variations in physicochemical characteristics, demands more emphasis on optimising the synthesis protocol for production of monodispersed nanozymes. In view of the discussed points it is evident that nanomedicine research arena is yet to fully mature to revolutionise the field of human medicine. High demand of investments, scientific and technical limits and overfilling sellers and marketing challenges such as excessive therapeutics prices and limited market penetration are obstacles that demand attention to enhance the potential of nanoparticle drug delivery share.
Development of novel delivery system for nanoencapsulation of catalase: formulation, characterization, and in vivo evaluation using oxidative skin injury model
Published in Artificial Cells, Nanomedicine, and Biotechnology, 2018
Heidi Mohamed Abdel-Mageed, Afaf S. Fahmy, Dalia S. Shaker, Saleh A. Mohamed
Michaelis–Menten constant (Km) and maximum reaction velocity (Vmax) for free CAT or e-CAT were calculated from activity assay using H2O2 as CAT substrate. The activity assay was performed using different concentrations of H2O2 ranging from 3 to 20 mM. The decomposition of each concentration of H2O2 was initiated at a fixed overall CAT concentration of 0.25 µg/ml. The catalytic efficiency (Vmax/Km) for both free CAT and e-CAT was also determined. CAT activity assay was carried out under standard assay conditions.