Synthesis and characterization of an N-acylsulfonamide inhibitor of human asparagine synthetase

Organic Letters

Volumn 5, Issue 12, 2003, Pages 2033-2036

  Koroniak, Lukasz ^a   Ciustea, Mihai ^a   Gutierrez, Jemy A ^a   Richards, Nigel G J ^a  

^a University of Florida ^*  (United States)

Author keywords

[No Author keywords available]

Indexed keywords

ASPARTATE AMMONIA LIGASE; ENZYME INHIBITOR; N ACYLSULFONAMIDE INHIBITOR; UNCLASSIFIED DRUG;

ARTICLE; DRUG SYNTHESIS; ENZYME ACTIVITY; ENZYME INHIBITION; HYDROGEN BOND; INHIBITION KINETICS; MOLECULAR INTERACTION;

ASPARTATE-AMMONIA LIGASE; ENZYME INHIBITORS; HUMANS; KINETICS; ORGANOPHOSPHORUS COMPOUNDS; RECOMBINANT PROTEINS; SULFONAMIDES;

EID: 0141742572     PISSN: 15237060     EISSN: None     Source Type: Journal    
DOI: 10.1021/ol034212n     Document Type: Article

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27. Briefly, the gene encoding human AS was obtained and modified so that the expressed enzyme would contain additional residues at the C-terminus corresponding to a c-myc tag. The resulting construct was then cloned into a pBAC-1 transfer vector (Novagen), which includes a C-terminal histidine tag for protein purification, so that expression of human AS is under control of the polh promoter allowing high levels of protein expression during late phases of viral infection. Recombination of pBAC-1 with the BacVector-3000 construct (Novagen) was then used successfully to obtain a baculovirus capable of replicating and infecting conditioned Sf9 cells, which also contained the gene encoding human AS. Using standard procedures, we expressed human AS in conditioned Sf9 cells, the recombinant enzyme being purified by chromatography on a preequilibrated Ni-Agarose column (see the Supporting Information). Additional details of this expression system can be found in the following references: (a) Farrell, P. J.; Lu, M. L.; Prevost, J.; Brown, C.; Behie, L.; Iatrou, K. Biotechnol. Bioeng. 1998, 60, 656-663. (b) Lenhard, T.; Reilander, H. Biochem. Biophys. Res. Commun. 1997, 238, 823-830.
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