Physics And Chemistry Basis Of Biotechnology - De Cuyper - tiera.ru

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Rational design of P450 enzymes for biotechnology content [77]. In order to circumvent this problem, in vitro investigation of the metabolic fate of potential drugs has been introduced and is currently being applied by the pharmaceutical industry at an early stage of trials of potential therapeutic agents. This is because the metabolic pattern of a drug influences its toxicological and therapeutic action. Thus decreased clearance of a candidate drug, due to being poorly metabolised by the host organism, will result to increased bioavailability and possibly toxicity. Investigation of the toxicity of the reactive metabolites formed is also very important. Finally, studying drug-drug interactions is highly necessary to predict the effect of P450 inhibition due to co-administration of an additional drug in multiple drug therapies [77]. Early investigation of the metabolic pattern of a candidate drug is mainly dictated by the need to reject potential drugs not fulfilling the pharmaceutical standards at the very initial stages of the trials, prior to clinical studies, for obvious financial and time-saving reasons. Different experimental systems for in vitro investigation of drug metabolism have been widely used, ranging from enzyme expression systems, to human hepatocytes and liver slices, as well as cell-free systems, such as liver homogenates and microsomes [79-80]. Each of these systems presents certain disadvantages and advantages, however, enzyme expression systems seem to be more widely accepted by the scientific community, as mimicking better the in vivo drug metabolism pathways and drug-drug interactions. Only a limited number of the P450s, present in the human liver, are involved in drug metabolism namely CYP 1A2, CYP 2C9, CYP 2C19, CYP 2D6 and CYP 3A4 [81-82]. However few others, including CYP 2E1, are reported to play a less significant role [83]. Due to the availability of the clone for the heterologous expression of CYP 2E1 in our laboratory, we focussed our studies on cytochrome CYP 2E1 [84]. CYP 2E1 is present in the liver and other tissues in many mammalian species. To date it has been shown to oxidise over 80 compounds including ethanol as well as carcinogens, environmental pollutants and drugs [85]. CYP 2E1 has been implicated to carcinogenesis, through the activation of procarcinogens, including potent tobaccospecific procarcinogens [86]. Finally the induction and stabilisation of CYP 2E 1 by ethanol consumption and the enzyme’s role in activating carcinogens is suspected to account for the increased carcinogenecity of certain compounds in alcoholics and this link has been shown in model systems for several compounds including bromobenzene, isoniazid and phenylbutazone [87]. 1.3. CHIMERAS OF P450 ENZYMES The production of catalytically self-sufficient P450 enzymes in bacterial hosts has a wide range of possible exciting applications. For instance cytochromes P450 metabolise a range of drugs and chemicals so bacteria containing these enzymes may help to further our understanding of human drug metabolism, the consequences of drug-drug interactions and the activation of carcinogens. The fusion protein hosts may also allow the mass production of compounds of interest by incorporating P450 biosynthetic pathways into one or a variety of hosts. Another application maybe the breakdown of environmental toxins, many of which are P450 substrates. The proteins may also further our understanding of the effect of P450 polymorphisms on human health by 79
Sheila J. Sadeghi et al incorporating various allelic variants into fusion protein systems and studying their different activities. 1.3.1. Bacterial P450-P450-reductase fusion protein systems So far only two naturally occurring catalytically self-sufficient bacterial P450s are known of the P450-P450-reductase fusion protein P450 BM3 from Bacillus megaterium and the fungal P450foxy from Fusarium oxysporum. De Montellano’s group [88] have also produced an artificial P450 from P450cam and its two reductase components putidaredoxin (Pd, a Fe-S protein) and putidaredoxin reductase (PdR, a FAD protein). P45Ocam was the first bacterial P450 to be crystallised and subsequently is the most studied and well understood of all P450s. Four fusion proteins were made by the same investigators, consisting of various combinations of the P450 and the reductase components. They found the PdR-Pd-P450cam construct to have the highest activity and also to be well coupled to oxygen consumption. It also had comparable activity to the reconstituted system at low protein concentrations (
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PHYSICS AND CHEMISTRY BASIS OF BIOT
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Physics and Chemistry Basis of Biot
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EDITORS PREFACE At the end of the 2
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TABLE OF CONTENTS EDITORS PREFACE .
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4.3. Expression and functionality .
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2 . Magnetically labelled antibodie
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6.1. Instrumental characterisation
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BIOMIMETIC MATERIALS SYNTHESIS Abst
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Biomimetic materials synthesis cont
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Biomimetic materials synthesis 3. I
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Biomimetic materials synthesis The
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Biomimetic materials synthesis 3.2.
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Biomimetic materials synthesis 3.5.
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Biomimetic materials synthesis In t
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Biomimetic materials synthesis The
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Biomimetic materials synthesis Othe
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Biomimetic materials synthesis on s
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Amperometric enzyme-based biosensor
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SUPPORTED LIPID MEMBRANES FOR RECON
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Supported lipid membranes for recon
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FUNCTIONAL STRUCTURE OF THE SECRETI
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Functional structure of the secreti
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COLD-ADAPTED ENZYMES D. GEORLETTE,
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Cold-adapted enzymes enzyme activit
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Cold-adapted enzymes their high spe
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Cold-adapted enzymes flexibility of
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Cold-adapted enzymes Some recent wo
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Cold-adapted enzymes Figure 2. Acti
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Cold-adapted enzymes is not complet
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Cold-adapted enzymes 16. Gilichinsk
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Cold-adapted enzymes 58. Rina, M.,
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Cold-adapted enzymes 101. Yip, K. S
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MOLECULAR AND CELLULAR MAGNETIC RES
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Molecular and cellular magnetic res
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RADIOACTIVE MICROSPHERES FOR MEDICA
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Radioactive microspheres for medica
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RADIATION-INDUCED BIORADICALS: Phys
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Radiation-induced bioradicals: phys
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RADIATION-INDUCED BIORADICALS: Tech
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Radiation-induced bioradicals: tech
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References Radiation-induced biorad
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AROMA MEASUREMENT: Recent developme
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Aroma measurement: recent developme
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62 63 64 65 66 67 68 69 70 71 72 73
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INDEX albumin.... 122,151, 198, 206
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frog embryo .......................
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P-32 ..............................
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