Catalyzes the hydrolysis of N-formyl-L-kynurenine to L-kynurenine, the second step in the kynurenine pathway of tryptophan degradation. Required for elimination of toxic metabolites
(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > cellular organisms: NE > Eukaryota: NE > Opisthokonta: NE > Metazoa: NE > Eumetazoa: NE > Bilateria: NE > Deuterostomia: NE > Chordata: NE > Craniata: NE > Vertebrata: NE > Gnathostomata: NE > Teleostomi: NE > Euteleostomi: NE > Sarcopterygii: NE > Dipnotetrapodomorpha: NE > Tetrapoda: NE > Amniota: NE > Mammalia: NE > Theria: NE > Eutheria: NE > Boreoeutheria: NE > Euarchontoglires: NE > Primates: NE > Haplorrhini: NE > Simiiformes: NE > Catarrhini: NE > Hominoidea: NE > Hominidae: NE > Homininae: NE > Homo: NE > Homo sapiens: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA MMDVSGVGFPSKVPWKKMSAEELENQYCPSRWVVRLGAEEALRTYSQIGI EATTRARATRKSLLHVPYGDGEGEKVDIYFPDESSEALPFFLFFHGGYWQ SGSKDESAFMVHPLTAQGVAVVIVAYGIAPKGTLDHMVDQVTRSVAFVQK RYPSNKGIYLCGHSAGAHLAAMMLLADWTKHGVTPNLRGFFLVSGVFDLE PIVYTSQNVALQLTLEDAQRNSPQLKVAQAQPVDPTCRVLVVVGQFDSPE FHRQSWEFYQTLCQGEWKASFEELHDVDHFEIVENLTQKDNVLTQIILKT IFQ
BACKGROUND: With the completion of the human genome sequence the functional analysis and characterization of the encoded proteins has become the next urging challenge in the post-genome era. The lack of comprehensive ORFeome resources has thus far hampered systematic applications by protein gain-of-function analysis. Gene and ORF coverage with full-length ORF clones thus needs to be extended. In combination with a unique and versatile cloning system, these will provide the tools for genome-wide systematic functional analyses, to achieve a deeper insight into complex biological processes. RESULTS: Here we describe the generation of a full-ORF clone resource of human genes applying the Gateway cloning technology (Invitrogen). A pipeline for efficient cloning and sequencing was developed and a sample tracking database was implemented to streamline the clone production process targeting more than 2,200 different ORFs. In addition, a robust cloning strategy was established, permitting the simultaneous generation of two clone variants that contain a particular ORF with as well as without a stop codon by the implementation of only one additional working step into the cloning procedure. Up to 92 % of the targeted ORFs were successfully amplified by PCR and more than 93 % of the amplicons successfully cloned. CONCLUSION: The German cDNA Consortium ORFeome resource currently consists of more than 3,800 sequence-verified entry clones representing ORFs, cloned with and without stop codon, for about 1,700 different gene loci. 177 splice variants were cloned representing 121 of these genes. The entry clones have been used to generate over 5,000 different expression constructs, providing the basis for functional profiling applications. As a member of the recently formed international ORFeome collaboration we substantially contribute to generating and providing a whole genome human ORFeome collection in a unique cloning system that is made freely available in the community.
The National Institutes of Health's Mammalian Gene Collection (MGC) project was designed to generate and sequence a publicly accessible cDNA resource containing a complete open reading frame (ORF) for every human and mouse gene. The project initially used a random strategy to select clones from a large number of cDNA libraries from diverse tissues. Candidate clones were chosen based on 5'-EST sequences, and then fully sequenced to high accuracy and analyzed by algorithms developed for this project. Currently, more than 11,000 human and 10,000 mouse genes are represented in MGC by at least one clone with a full ORF. The random selection approach is now reaching a saturation point, and a transition to protocols targeted at the missing transcripts is now required to complete the mouse and human collections. Comparison of the sequence of the MGC clones to reference genome sequences reveals that most cDNA clones are of very high sequence quality, although it is likely that some cDNAs may carry missense variants as a consequence of experimental artifact, such as PCR, cloning, or reverse transcriptase errors. Recently, a rat cDNA component was added to the project, and ongoing frog (Xenopus) and zebrafish (Danio) cDNA projects were expanded to take advantage of the high-throughput MGC pipeline.
Title: Kynurenine formamidase: determination of primary structure and modeling-based prediction of tertiary structure and catalytic triad Pabarcus MK, Casida JE Ref: Biochimica & Biophysica Acta, 1596:201, 2002 : PubMed
Kynurenine formamidase (KFase) (EC 3.5.1.9) hydrolyzes N-formyl-L-kynurenine, an obligatory step in the conversion of tryptophan to nicotinic acid. Low KFase activity in chicken embryos, from inhibition by organophosphorus insecticides and their metabolites such as diazoxon, leads to marked developmental abnormalities. While KFase was purportedly isolated previously, the structure and residues important for catalysis and inhibition were not established. KFase was isolated here from mouse liver cytosol by (NH4)2SO4 precipitation and three FPLC steps (resulting in 221-fold increase in specific activity for N-formyl-L-kynurenine hydrolysis) followed by conversion to [3H]diethylphosphoryl-KFase and finally isolation by C4 reverse-phase high-performance liquid chromatography. Determination of tryptic fragment amino acid sequences and cDNA cloning produced a new 305-amino-acid protein sequence. Although an amidase by function, the primary structure of KFase lacks the amidase signature sequence and is more similar to esterases and lipases. Sequence profile analysis indicates KFase is related to the esterase/lipase/thioesterase family containing the conserved active-site serine sequence GXSXG. The alpha/beta-hydrolase fold is suggested for KFase by its primary sequence and predicted secondary conformation. A three-dimensional model based on the structures of homologous carboxylesterase EST2 and brefeldin A esterase implicates Ser162, Asp247 and His279 as the active site triad.
Homo sapiens (Human) esterase D (EC 3.1.1.1) formylglutathione hydrolase