Directed enzymes

Directed Enzyme Evolution Screening and Selection Methods, Humana Press, Totowa. Vol. 230. (b) Brakmann, S. and Johnsson, K. (eds)(2002) Directed Molecular Evolution of Proteins (or How to Improve Enzymes for Biocatalysis), Wdey-VCH Verlag GmbH, Weinheim. (c) Brakmann, S. and Schwienhorst, A. (eds)... 

The improvement in the enantioselectivity from 28% (P) to >99% (S) was due to the suppression of every enzyme that reduces the substrate, followed by the stimulation of an S-directing enzyme by the addition of the coenzyme and an excess amount of 2-propanol, agents which push the equilibrium toward the reduction of the substrate. 

It was confirmed, by separating the enzymes in the powder, that many S- and R-directing enzymes do indeed exist in the dried cells. The addition of a coenzyme and cyclopentanol stimulates only an S enzyme because the specific S enzyme can oxidize cyclopentanol (concomitantly reducing NAD(P) ), while other S or R enzymes cannot use cyclopentanol as effectively [14cj. This presents a very interesting case, where the experimental conditions of reduction with a cell having both S-and R-directing enzymes was modified and resulted in excellent S enantioselectivity. 

VI. Conformationally Restricted Active-Site-Directed Enzyme Inhibitors... 

VI. CONFORMATIONALLY RESTRICTED ACTIVE-SITE-DIRECTED ENZYME INHIBITORS... 

Fisher, J. Ramakrishnan, K. Becvar, J. E. Direct enzyme-catalyzed reduction of anthra-cyclines by reduced nicotinamide adenine dinucleotide. Biochemistry 1983, 22, 1347-1355. 

Directed Enzyme Evolution and High-Throughput Screening... 

Arnold, F.H. and Georgiou, G. (eds) (2003) Directed Enzyme Evolution Screening and Selection Methods, Methods in Molecular Biology, vol. 230, Humana Press, Inc., Clifton, NJ. 

Farinas, E.T., Bulter, T., Arnold, F.H., Directed enzyme evolution, Curr. Opin. Biotech-nol. 2001, 22, 545-551... 

Very limited information is available through direct enzyme studies for DszA. However, its specificity for couple of possible intermediates resulting from DszC reaction has been studied. The DszA enzyme was capable of converting dibenz[c,e][l,2]oxanthiin 6,6-dioxide (sultone) as well as sultine to 2,2 dihydoxybiphenyl (DHBP), in addition to its inherent activity towards DBT sulfone. However, the relative activities were lower than the activity towards DBT sulfone (54% and 23%, respectively). The proposed pathway for this reaction is given in Fig. 2 [26,153],... 

Structure 139 represents a cephalosporin conjugate of SIN-1 13 that has been synthesized and evaluated as a /3-lactamase-dependent, nitric oxide-releasing conjugate with potential application in antibody-directed enzyme prodrug therapy (ADEPT) <2003BML1687>. 

Figure 13.7 Outline of antibody-directed enzyme prodrug therapy (ADEPT). Subsequent to its enzymatic activation, the active drug is taken up by the cell, upon which it exhibits a cytocidal effect. Refer to text for specific detail...

A review describes how penicillins and cephalosporins having S-aminosulfenamine side-chains at the 6- and 7-positions, respectively, may act as P-lactamase-dependent prodrugs either as antibiotics or in antibody-directed enzyme prodmg therapy (ADEPT) in the treatment of cancer <00T5699>. 

Fig. 6.14. Schematic representation of selective delivery obtained by gene-directed enzyme-prodrug therapy (GDEPT). The gene encoding an exogenous enzyme is transferred to tumor cells, where it is to be expressed. In a second step, a prodrug is administered that is selectively activated at the tumor site by the exogenous enzyme expressed by the tumor cells.

W. A. Denny, W. R. Wilson, The Design of Selectively-Activated Anti-Cancer Prodrugs for Use in Antibody-Directed and Gene-Directed Enzyme Prodrug Therapies , J. Pharm. Pharmacol. 1998, 50, 387 - 394. 

I. Niculescu-Duvaz, F. Friedlos, D. Niculescu-Duvaz, L. Davies, C. J. Springer, Prodrugs for Antibody- and Gene-Directed Enzyme Prodrug Therapies (ADEPT and GDEPT) , Anti-Cancer Desing 1999,14, 517-538. 

M. P. Napier, S. K. Sharma, C. J. Springer, K. D. Bagshawe, A. J. Green, J. Martin, S. M. Stribbling, N. Cushen, D. O Malley, R. H. Begent, Antibody-Directed Enzyme Prodrug Therapy Efficacy and Mechanism of Action in Colorectal Carcinoma , Clin. Cancer Res. 2000, 6, 765-772. 

G. K. Smith, S. Banks, T. A. Blumenkopf, M. Cory, J. Humphreys, R. M. Laethem, J. Miller, C. P. Moxham, R. Muhin, P. H. Ray, L. M. Walton, L. A. Wolfe III, Toward Antibody-Directed Enzyme Prodrug Therapy with the T286G Mutant of Human Carboxypeptidase A1 and Novel in vivo Stable Prodrugs of Methotrexate , J. Biol. Chem. 1997, 272, 15804-15816. 

K. D. Bagshawe, Antibody Directed Enzymes Revive Anti-Cancer Prodrugs Concept , Br. J. Cancer 1987, 56, 531-532 K. D. Bagshawe, Antibody-Directed Enzyme/Pro-drug Therapy (ADEPT) , Biochem. Soc. Trans. 1990, 18, 750-752. 

A. B. Mauger, P. J. Burke, H. H. Somani, F. Friedlos, R. J. Knox, Self-Immolative Prodrugs Candidates for Antibody-Directed Enzyme Prodrug Therapy in Conjunction with a Nitroreductase Enzyme , J. Med. Chem. 1994, 37, 3452-3458. 

J. Cassidy, Polymer-Directed Enzyme Prodrug Therapy , Drug News Perspect. 2000, 13, 477 - 480. 

P. H. J. Houba, R. G. G. Leenders, E. Boven, J. W. Scheeren, H. M. Pinedo, H. J. Hais-ma, Characterization of Novel Anthracycbne Prodrugs Activated by Human /3-Glucuronidase for Use in Antibody-Directed Enzyme Prodrug Therapy , Biochem. Pharmacol. 

Y. L. Leu, S. R. Roffler, J. W. Chern, Design and Synthesis of Water-Soluble Glucuro-nide Derivatives of Campothecin for Cancer Prodrug Monotherapy and Antibody-Directed Enzyme Prodrug Therapy (ADEPT) , J. Med. Chem. 1999, 42, 3623 - 3628. 

Gupta, R.D. and Tawfik, D.S., Directed enzyme evolution via small and effective neutral drift libraries. Nat. Methods, 2008, 5, 939-942. 

The thiosulfate reductase/rhodanese/APS reductase system is thus supported by evidence from direct enzyme assay, whole-cell metabolism and energetics, and S-labehng experiments and provides a robust hypothesis to explain thionate oxidation and energy conservation in at least some chemolithotrophs. 

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