BIOCHEMICAL AND MOLECULAR GENETIC STUDIES OF UROPORPHYRINOGEN III SYNTHASE.
Item
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Title
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BIOCHEMICAL AND MOLECULAR GENETIC STUDIES OF UROPORPHYRINOGEN III SYNTHASE.
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Identifier
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AAI8801771
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identifier
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8801771
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Creator
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TSAI, SHIH-FENG.
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Contributor
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Robert Desnick
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Date
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1987
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Genetics
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Abstract
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Uroporphyrinogen III synthase (URO-S) catalyzes the formation of the cyclic tetrapyrrole, uroporphyrinogen (URO'gen) III, from the linear tetrapyrrole, hydroxymethylbilane (HMB). The deficiency of URO-S is the biochemical defect in congenital erythropoietic porphyria (CEP). URO-S from human erythrocytes has been purified to homogeneity (Tsai et al., J. Biol. Chem. 262:1268-1273, 1987) and, amino acid sequences for the N-terminal and tryptic peptides have been determined. Synthetic oligonucleotide mixtures were used to screen a human adult liver cDNA library. Eight positive clones were isolated from a total of 1.2 {dollar}\times{dollar} 10{dollar}\sp6{dollar} recombinants screened. Of these, one clone designated pURO-S 2, had an insert of {dollar}\cong{dollar}1.3 kb which contained 5{dollar}\sp\prime{dollar} and 3{dollar}\sp\prime{dollar} untranslated sequences of 196 and 284 bp, respectively. An open reading frame of 798 bp was identified, which encoded a protein of 265 amino acids with a molecular weight of 28,607. The authenticity of this clone was confirmed by the colinearity of the predicted amino acid sequence with 81 residues determined by microsequencing the purified enzyme. Northern hybridization of poly(A{dollar}\sp{lcub}+{rcub}{dollar}) RNA revealed a 1.3 kb transcript in normal human cultured lymphoblasts. The molecular cloning and sequencing of a full-length cDNA for URO-S should facilitate the studies of the structure, organization and expression of this human heme biosynthetic gene, as well as the molecular nature of the lesions in CEP.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biomedical Sciences
