Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.6.1.44 (AGT)
 770 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously detected a single base substitution of G by A at the Arg codon CGC in exon 4 of the mutant lactate dehydrogenase (LDH) gene, an unstable LDH-B variant (case 1). Here, we use the polymerase chain reaction (PCR) to amplify genomic DNA of two cases (the original case 1 and a new patient, case 2). We were able to confirm that case 1 is homozygous for the mutation, causing a replacement of the conserved Arg by His at residue 173. The resulting LDH-B variant subunit is unstable in vivo. Whereas the mutation in exon 4 was not observed in case 2, a different single base substitution of A by C was detected at the Ser codon AGT in exon 3. This mutation causes a replacement of the conserved Ser by Arg at residue 131. Genomic analysis of the family of case 2 by mismatched PCR showed that the missense mutation was consistent with their biochemical phenotypes. The replacement results in a conformational change of the residues near the Ser, probably because the side chain of Arg is much more bulky than that of Ser. The change may affect the arrangement of the cofactor binding site and result in the loss of enzyme activity. The experimental observations are consistent with computer graphics analyses.
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PMID:Molecular characterization of genetic mutations in human lactate dehydrogenase (LDH) B (H) variant. 158 25

Hereditary elliptocytosis (HE) Sp alpha I/74 is a disorder associated with defective spectrin (Sp) heterodimer self-association and an abnormal tryptic cleavage of the 80-kD alpha I domain of Sp resulting in increased amounts of a 74-kD peptide. The molecular basis of this disorder is heterogeneous and mutations in codons 28, 46, 48, and 49 (codons 22, 40, 42, and 43 in the previous nomenclature which did not include the six NH2-terminal amino acids) have been reported. In this study we present data on seven unrelated HE Sp alpha I/74 kindred from diverse racial backgrounds in whom we identified four different mutations all occurring in exon 2 of alpha Sp at codon 28. Utilizing the polymerase chain reaction we established a CGT----CTT; Arg----Leu 28 mutation in one kindred of Arab/Druze origin. In two unrelated white kindred of English/European origin the substitution is CGT----AGT; Arg----Ser 28 and in two apparently unrelated white kindred from New Zealand, the mutation is CGT----TGT; Arg----Cys 28. Finally, in one American black kindred and in a black kindred from Ghana the mutation involves CGT----CAT; Arg----His 28. Allele specific oligonucleotide hybridization confirmed that the probands are heterozygous for the respective mutant alleles. All four point mutations abolished an Aha II restriction enzyme site which allowed verification of linkage of the mutation with HE Sp alpha I/74. Our results imply that codon 28 of alpha Sp is a "hot spot" for mutations and also indicate that Arg 28 is critical for the conformational stability and functional self association of Sp heterodimers.
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PMID:Four different mutations in codon 28 of alpha spectrin are associated with structurally and functionally abnormal spectrin alpha I/74 in hereditary elliptocytosis. 167 39

Hereditary pyropoikilocytosis (HPP) and hereditary elliptocytosis are closely related, congenital disorders of the red blood cell usually associated with defective spectrin self-association and abnormal limited tryptic digestion of the N-terminal of domain of spectrin. Enhanced cleavage by trypsin of spectrin from affected individuals at arginyl residue 45* and lysyl residue 48* frequently yields increased amounts of an alpha 1/74-Kd fragment at the expense of the normal alpha 1/80-Kd parent fragment. Limited tryptic digestion of three unrelated individuals with HPP showed the alpha 1/74 defect. To ascertain the molecular defect responsible for the abnormality, the structure of exon 2 of the alpha-spectrin gene was examined. Genomic DNA from the subjects was amplified by the polymerase chain reaction using primers flanking exon 2. Restriction endonuclease digestion of amplified products showed the loss of the HindIII site at codons 47 and 48 in one allele of subject 1 and abolished the AhaII site at codons 27 and 28 in one allele of subjects 2 and 3. Nucleotide sequence analysis of subcloned amplified DNA from the HPP subjects showed three novel amino acid substitutions. In subject 1 (a black individual), a single base substitution (AAG----AGG) at codon position 48 changes amino acid residue lysine to arginine. In subject 2 (a white individual), a single base substitution (CGT----AGT) at codon 28 changes arginine to serine. In subject 3 (a black individual), a different base substitution at position 28 (CGT----CTT) changes arginine to leucine. These mutations occur at positions of the alpha l domain where other mutations have also been described, indicating that the normal residues at these positions play an important role in spectrin dimer self-association and thus, in membrane stability.
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PMID:Heterogeneity of the molecular basis of hereditary pyropoikilocytosis and hereditary elliptocytosis associated with increased levels of the spectrin alpha I/74-kilodalton tryptic peptide. 187 97

Dimethylarginine:pyruvate aminotransferase, which plays a role in the metabolism of dimethylarginines, has been purified to homogeneity from rat kidney. The enzyme has a molecular weight of approximately 200,000 and an isoelectric point at about pH 6.3. The enzyme consists of four similar subunits having a molecular weight of about 50,000. The enzyme catalyzes the effective transaminations of guanidino-N methylated L-arginines (e.g. NG,NG-dimethyl-L-arginine, NG,N'G-dimethyl-L-arginine and NG-monomethyl-L-arginine) and the alpha-amino group of L-ornithine to pyruvate or glyoxylate. The enzyme was always accompanied by the known alanine:glyoxylate amino-transferase activity with the ratios of their specific activities remaining constant during the purification steps. The physicochemical and immunological properties of the purified enzyme were shown to be identical with those of the isozyme of alanine:glyoxylate aminotransferase (EC 2.6.1.44), designated as alanine:glyoxylate aminotransferase 2 (Noguchi, T. (1987) in Peroxisomes in Biology and Medicine (Fahimi, H. D., and Sies, H., eds) pp. 234-243, Springer-Verlag, Heidelberg). The distribution profiles in tissues and the negative response to glucagon treatment further supported the identity of the two enzymes. The present data show that alanine:glyoxilate aminotransferase 2 functions in dimethylarginine metabolism in vivo in rats.
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PMID:Dimethylarginine:pyruvate aminotransferase in rats. Purification, properties, and identity with alanine:glyoxylate aminotransferase 2. 212 86

Limited tryptic digestion of spectrin (Sp) from seven related individuals manifesting hereditary elliptocytosis (HE) or hereditary pyropoikilocytosis (HPP) phenotypes revealed the presence of a novel peptide with a molecular weight of 78 Kd and a concomitant decrease in the alpha I domain (80-Kd peptide), which is the domain involved in the dimer self-association process. Sp from the normal members of this white family exhibited a normal peptide pattern, as compared with controls. The abnormal peptide pattern was associated with a decreased ability of Sp dimer to self-associate. In this kindred in which three generations were available for study, the clinical manifestations were quite variable and ranged from the asymptomatic HE carrier state to hemolytic HE or to severe anemia requiring splenectomy. The severity of the disease appeared to be correlated both with the amount of mutant spectrin (31% to 69%) and with the excess of the Sp dimer found in the membrane (26% to 60%, compared with a normal value of 5.6% +/- 2.2%). Partial amino acid sequencing showed that the alpha I/78-Kd peptide resulted from cleavage at lysine residue 10 of the alpha I/80-Kd domain. Knowledge of the exon/intron structure of cloned genomic DNA encoding the alpha I domain allowed us to amplify in vitro a DNA fragment containing the third exon of the alpha-spectrin gene. The amplified fragment was subcloned and sequenced. A G to T transversion was found in the 39th codon (AGT for AGG), which changed the normal arginine to a serine. Hybridization of amplified DNAs with allele-specific oligonucleotides corresponding to the normal and mutant sequences confirmed the presence of the mutation in three other HE members of the family (the propositus mother, brother, and sister).
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PMID:Sp alpha I/78: a mutation of the alpha I spectrin domain in a white kindred with HE and HPP phenotypes. 256 62

Two sisters presented with severe insulin resistance and markedly decreased insulin binding to erythrocytes, cultured fibroblasts and transformed lymphocytes. The dose-response curve of insulin-stimulated amino acid uptake in the fibroblasts was shifted to the right. The molecular weight of the insulin receptor on the transformed lymphocytes from the patients was 210,000 and could not be dissociated to alpha- and beta-subunits by dithiothreitol treatment. However, the proreceptor was cleaved by trypsin and this led to the production of alpha-subunit with normal insulin binding. We performed cDNA sequence analysis of the cleavage site of the insulin proreceptor from the patients. The polymerase chain reaction was used to obtain a large amount of cDNA coding for the region including the interconnecting site. A thermostable DNA polymerase, Taq polymerase, successfully produced enough cDNA for the region to be sequenced. The results showed an AGG (Arg) to AGT (Ser) point mutation, resulting in the change of the interconnecting sequence of the two subunits from -Arg-Lys-Arg-Arg- to -Arg-Lys-Arg-Ser-. These results suggest that the tertiary structure change of the cleavage site leads to production of unprocessed insulin proreceptors.
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PMID:Unprocessed insulin proreceptors due to point mutation at the cleavage site. 268 Mar 65

In the accompanying paper, we present and analyse the sequence of a "superactivator" mutant allele of the CYP1 (HAP1) gene. This locus encodes a trans-acting pleiotropic positive regulator of the transcription of both isocytochrome c structural genes. In this paper, we present the genetic localization of the mutation and the sequence of the wild-type fragment that includes the mutation. The mutated phenotype that commutes the expression of the two isocytochrome structural genes (superactivation of CYP3 and inhibition of CYC1) results from a transversion in an AGT codon (serine) in the wild-type to an AGG codon (arginine) in the mutant. Moreover, we show that the missense mutation that affects the amino acid preceding the first cysteine of the "Zn finger" is responsible on its own account for the entire mutated phenotype. In all seven yeast regulatory proteins analysed so far, this position is occupied by a neutral amino acid (serine, alanine or glycine), thus the serine-arginine replacement is a radical one. This result is consistent with the hypothesis of alternative and mutually exclusive Zn fingers, formed either at low or high redox potential, recognizing the target sequences identified in the upstream regions of the CYC1 and CYP3 isocytochrome c structural genes.
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PMID:CYP1 (HAP1) regulator of oxygen-dependent gene expression in yeast. II. Missense mutation suggests alternative Zn fingers as discriminating agents of gene control. 285 59

Failure to cleave the interconnecting site between alpha- and beta-subunit produced insulin proreceptors in the plasma membranes which had markedly low affinity to insulin, leading to extreme insulin resistance in a patient. We performed cDNA sequence analysis of the cleavage site of the insulin proreceptor from the patient. Polymerase chain reaction was used to obtain large amount of cDNA coding for the region including the interconnecting site. A thermostable DNA polymerase, Taq polymerase, successfully produced enough amount of cDNA of the region to be sequenced. The results showed AGG (Arg) to AGT (Ser) point mutation, resulting in the change of interconnecting sequence of the two subunits from -Arg-Lys-Arg-Arg- to -Arg-Lys-Arg-Ser-. These results suggest that the tertial structure change of the cleavage site leads to production of unprocessed insulin proreceptors.
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PMID:Insulin resistance by unprocessed insulin proreceptors point mutation at the cleavage site. 328 35

In order to confirm the amino acid sequence predicted from the nucleotide sequence of cDNA and also to elucidate the intracellular localization and molecular evolution, human liver alanine-glyoxylate transaminase 1 (AGT1) was purified and subjected to partial amino acid sequence determination, with special attention to posttranslational modification. The enzyme was purified to homogeneity from the 10,000 x g supernatant of human liver homogenate. The purified enzyme showed only a single protein band at about 43 kDa on SDS-PAGE, indicating that it is a homodimer of two identical subunits, because the native enzyme has a molecular mass of about 80 kDa. Both the amino- and carboxyl-terminal peptides of the enzyme were isolated from a cyanogen bromide digest of the S-carboxyl-methylated protein and subjected to amino acid sequence determination. The alpha-amino group of the amino-terminal peptide was shown to be blocked by an acetyl group. The carboxyl-terminal sequence contained a putative N-glycosylation sequence (-Asn-Ala-Thr-), the only one present in the whole molecule, but this sequence was normally determined, indicating that the enzyme is not N-glycosylated. Purdue et al. [J. Cell Biol. 111, 2341-2351 (1990)] have reported that Pro-11, Gly-170, and Ile-340 in normal human AGT1 were replaced by Leu, Arg, and Met, respectively, in a patient with primary hyperoxaluria type 1. We confirmed that residue-11 was Pro. Both the amino- and carboxyl-terminal sequences of the enzyme showed extensive similarity with those of rat liver mitochondrial serine-pyruvate aminotransferase and the small chain of hydrogenase from a thermophilic unicellular cyanobacterium, Synechococcus PCC 6716.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Purification and amino- and carboxyl-terminal amino acid sequences of alanine-glyoxylate transaminase 1 from human liver. 779 68

We have evidence that the limulus (Tachypleus tri-dentatus) hemocyte transglutaminase (TGase) has a molecular mass of 86 kDa and properties of the mammalian type II TGase-like enzyme (Tokunaga, F., Yamada, M., Miyata, T., Ding, Y.-L., Hiranaga-Kawabata, M., Muta, T., Iwanaga, S., Ichinose, A., and Davie, E.W. (1993) J. Biol. Chem. 268, 252-261). We present here the cDNA and amino acid sequences, and localization of the TGase in various tissues of limulus. The cloned cDNA for TGase consists of 2,884 base pairs. An open reading frame of 2,292 base pairs encodes a sequence comprising 764 residues of the mature protein with molecular masses of 87,021 and 87,110 Da, due to two different clones. The discrepancies of nucleotides in these two clones result in 3 amino acid exchanges at positions Gly452(GGT)-Arg(CGT), Ser477(AGT)-Cys(TGT), and Ile486(ATC)-Ser(AGC), respectively. Northern blot analysis on a total RNA extracted from various tissues of limulus revealed that TGase is expressed with 3.0 kilobases of a single type of mRNA, mainly in hemocytes, hepatopancreas, and gastric tissues. Limulus TGase shows significant sequence similarity with the mammalian TGase family, as follows: guinea pig liver TGase (32.7%), human factor XIIIa subunit (34.7%), human keratinocyte TGase (37.6%), and human erythrocyte band 4.2 (23.0%). Limulus TGase has a unique NH2-terminal cationic extension of 60 residues with no homology to the NH2 termini of mammalian TGases. Based on the alignment of the amino acid sequence of limulus TGase with those of the known TGase family, a phylogenetic tree representing an evolutionary relationship among the family members was inferred by the neighbor joining method.
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PMID:Limulus hemocyte transglutaminase. cDNA cloning, amino acid sequence, and tissue localization. 809 43


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