Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Y1 mouse adrenocortical tumor cell line retains the ability to synthesize and secrete steroids, but does not express steroid 21-hydroxylase (C21) and, therefore, does not produce 21-hydroxylated steroids. In this investigation the mechanisms underlying the loss of C21 activity in the Y1 cell line were explored. A 9-kilobase BglII fragment containing the C21 gene was cloned from the Y1 genome. This genomic clone directed the synthesis of C21 transcripts and 21-hydroxylated steroid products when transfected back into the Y1 cell line. As determined by restriction endonuclease digestions with MspI and HpaII, enzymes that distinguish between unmethylated and methylated CCGG sites, the endogenous C21 gene was extensively methylated in Y1 adrenal cells and in cells from other mouse tissues that do not normally express this gene. In contrast, the C21 gene was hypomethylated in primary cultures of mouse adrenal cells which normally synthesize large amounts of C21. The cloned C21 gene transfected into Y1 cells initially was unmethylated, but became extensively methylated with prolonged culture of the cells; prolonged culture of these transfectants also resulted in a loss of C21 expression. Loss of C21 expression in Y1 transfectants, however, temporally preceded the extensive methylation of the transfected C21 gene. Furthermore, treatment of Y1 cells with 5-azacytidine caused a demethylation of the endogenous C21 gene, but did not result in the recovery of C21 expression. These results indicate that Y1 cells contain a functional C21 gene that has been silenced by a reversible cis-modification event.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:cis modification of the steroid 21-hydroxylase gene prevents its expression in the Y1 mouse adrenocortical tumor cell line. 170 55

A point mutation within exon 7 producing an amino acid coding change and a recognition site for the endonuclease Ncol has been reported in the HLA-Bw47-linked CYP21A pseudogene and some mutant CYP21B (steroid 21-hydroxylase) genes of patients with congenital adrenal hyperplasia (CAH). Whether this mutation is deleterious was not demonstrated. We analyzed DNA from various subjects for the presence of the exon 7 Ncol site: group 1, 10 normal subjects; group 2, 11 patients with salt-losing CAH; and group 3, 18 members of an Amish pedigree in which 10 expressed HLA-Bw47 not linked to CAH. Southern blots of Ncol-digested genomic DNA which were hybridized with CYP21 cDNA showed that four subjects of group 1 had a heterozygous Ncol pattern. In group 2, seven patients had the Ncol site; two of them were homozygous for the site and had deletions of both CYP21B genes. The other five were heterozygous for the Ncol site, which was linked to a CYP21B deletion and a HLA-Bw47 haplotype. In group 3, no one exhibited the exon 7 Ncol site. To map the Ncol sites to CYP21A or CYP21B in the normal subjects, DNA from the four Ncol heterozygous subjects was double digested with Ncol and Mbol and hybridized with CYP21 cDNA. Ncol-Mbol fragments unique to CYP21A were identified in all four, but the smaller CYP21B-specific fragments were not detected. Their genomic DNA in the region of exon 7 (bases +1167 to +2058) was then amplified, cloned, and sequenced.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Exon 7 Ncol restriction site within CYP21B (steroid 21-hydroxylase) is a normal polymorphism. 197 47

Nonclassic steroid 21-hydroxylase deficiency is a frequent, relatively mild disorder of cortisol biosynthesis characterized by variable signs of postnatal androgen excess. It is inherited as an allelic variant of the CYP21B gene encoding the 21-hydroxylase enzyme. CYP21B is located in the HLA histocompatibility complex, and a nonclassic allele is often associated with characteristic HLA antigens: B14;DR1. A CYP21B gene from a HLA-B14;DR1 homozygous patient with nonclassic 21-hydroxylase deficiency was cloned and analyzed. Five deviations from the normal sequence of CYP21B were found, but only one appeared likely to affect the functional integrity of the protein: codon 281, GTG, encoding valine, was changed to TTG, leucine. An oligonucleotide probe was constructed corresponding to the mutant sequence surrounding codon 281 and hybridized with DNA samples digested with restriction endonuclease Taq I. Samples from 8 nonclassic 21-hydroxylase deficiency patients carrying HLA-B14;DR1 contained a hybridizing fragment 3700 base-pairs long, indicating presence of the val-281 mutation in the CYP21B gene. In contrast, unaffected individuals and one patient who lacked HLA-B14;DR1 showed no evidence of the val-281 mutation in CYP21B. We conclude that the codon 281 mutation is a consistent molecular genetic marker for nonclassic 21-hydroxylase deficiency associated with HLA-B14;DR1.
...
PMID:Clinical and genetic characterization of nonclassic 21-hydroxylase deficiency. 278 81

Nonclassic steroid 21-hydroxylase deficiency is a frequent, relatively mild disorder of cortisol biosynthesis, characterized by variable signs of postnatal androgen excess. It is inherited as an allelic variant of the gene designated CYP21B, which encodes 21-hydroxylase. CYP21B is located in the HLA histocompatibility complex, and a "nonclassic" allelic variant is often associated with characteristic HLA antigens--B14,DR1. We cloned and analyzed the CYP21B gene from a patient homozygous for HLA-B14,DR1 who had nonclassic 21-hydroxylase deficiency. Five deviations from the normal genetic sequence of CYP21B were found, but only one appeared likely to affect the functional integrity of the protein: codon 281, GTG, encoding valine, was changed to TTG, leucine. We constructed an oligonucleotide probe corresponding to the mutant DNA sequence surrounding codon 281 and hybridized the probe with DNA samples digested with the restriction endonuclease Taql. Samples from eight patients with nonclassic 21-hydroxylase deficiency who had the haplotype HLA-B14,DR1 contained a hybridizing fragment 3700 base pairs long, indicating the presence of the valine-281 mutation in the CYP21B gene. In contrast, unaffected subjects and one patient with nonclassic deficiency who did not have HLA-B14,DR1 had no evidence of this mutation. We conclude that the mutation in codon 281 is a consistent molecular genetic marker for nonclassic 21-hydroxylase deficiency associated with HLA-B14,DR1.
...
PMID:Molecular genetic analysis of nonclassic steroid 21-hydroxylase deficiency associated with HLA-B14,DR1. 326 7

Congenital adrenal hyperplasia (CAH) is a common genetic disorder due to defective 21-hydroxylation of steroid hormones. The human P450XXIA2 gene encodes cytochrome P450c21 [steroid 21-monooxygenase (steroid 21-hydroxylase), EC 1.14.99.10], which mediates 21-hydroxylation. The P450XXIA2 gene may be distinguished from the duplicated P450XXIA1 pseudogene by cleavage with the restriction endonuclease Taq I, with the XXIA2 gene characterized by a 3.7-kilobase (kb) fragment and the XXIA1 pseudogene characterized by a 3.2-kb fragment. Restriction endonuclease mapping by several laboratories has suggested that deletion of the P450XXIA2 gene occurs in about 25% of patients with CAH, as their genomic DNA lacks detectable 3.7-kb Taq I fragments. We have cloned human P450c21 cDNA and used it to study genomic DNA prepared from 51 persons in 10 families, each of which includes 2 or more persons with CAH. After Taq I digestion, apparent deletions are seen in 7 of the 20 alleles of the probands; using EcoRI, apparent deletions are seen in 9 of the 20 alleles. However, the apparently deleted alleles seen with Taq I do not coincide with those seen with EcoRI. Furthermore, studies with Bgl II, EcoRI, Kpn I, and Xba I yield normal patterns with at least two enzymes in all cases. Since all probands yielded normal patterns with at least two of the five enzymes used, we conclude that the P450XXIA2 gene "deletions" widely reported in CAH patients probably represent gene conversions, unequal crossovers, or polymorphisms rather than simple gene deletions.
...
PMID:P450XXI (steroid 21-hydroxylase) gene deletions are not found in family studies of congenital adrenal hyperplasia. 349 99

Genomic DNAs from twelve Japanese patients with steroid 21-hydroxylase [21-OHase; steroid 21-monooxygenase; steroid, hydrogen-donor:oxygen oxidoreductase (21-hydroxylating); EC 1.14.99.10] deficiency were analyzed by Southern blot hybridization. A 3.7-kilobase (kb) Taq I and a 1.7-kb Pvu II restriction endonuclease fragment that correspond to a 21-OHase B gene were absent from the DNA of two unrelated patients with the salt-wasting form of the disease. However, a 10.5-kb Bgl II fragment corresponding to the region encompassing the 21-OHase B gene was still present in these two patients. The genes encoding 21-OHase were cloned from one of these two patients, who was homozygous by descent for HLA-A26;B39;C4A3;C4B1;DR4. Restriction endonuclease mapping as well as partial nucleotide sequencing analysis revealed that the 21-OHase B gene of the patient has been converted to the pseudogene, 21-OHase A, as far as the critical 0.5-kb sequence was concerned. Thus, the defect was due to both chromosomes each carrying two copies of 21-OHase A pseudogene and lacking functional 21-OHase B gene.
...
PMID:Gene conversion-like events cause steroid 21-hydroxylase deficiency in congenital adrenal hyperplasia. 350 Apr 73

A search for uncharacterized genes of the S region of the murine H-2 major histocompatibility complex was undertaken; a series of cosmid clones previously aligned by overlap hybridizations were used as radiolabeled probes. Sequences hybridizing with liver poly(A)+ RNA were found within a cosmid covering a region 3' to the C4-Slp gene (the gene encoding the hemolytically inactive isoform of the fourth component of serum complement). Radiolabeled, short cDNA complementary to liver poly(A)+ RNA was used to establish the transcriptional polarity of the newly detected gene and to define fragments containing its 3' end. DNA sequence analyses and comparisons with porcine peptides established that the gene encodes the enzyme steroid 21-hydroxylase (EC 1.14.99.10), a cytochrome P-450 often referred to as P-450(C21), whose major site of expression is the adrenal gland. Two copies of the P-450(C21) gene, very similar yet distinguishable by restriction endonuclease analysis, were found individually associated with C4 and C4-Slp, genes that encode isoforms of mouse fourth component of complement. One of the P-450(C21) genes is coamplified with C4-Slp in H-2w7, a haplotype carrying a rare elongation of the S region. Comparisons with other members of the P-450 gene family show that the P-450(C21) genes encode peptides of extraordinary evolutionary conservation. The detection of a liver transcript of P-450(C21) raises the issue of the specific metabolic role of this enzyme in this organ and may have implications for the interpretation of human congenital adrenal hyperplasia.
...
PMID:Liver mRNA probes disclose two cytochrome P-450 genes duplicated in tandem with the complement C4 loci of the mouse H-2S region. 387 1

Many human diseases are caused by small alterations in the genes and in the majority of cases sophisticated protocols are required for their detection. In this study we estimated the efficacy of an enzymatic protocol, which using a new mismatch-specific DNA plant endonuclease from celery (CEL family) recognizes and cleaves mismatched alleles between mutant and normal PCR products. The protocol was standardized on a variety of known mutations, in 11 patients with cystic fibrosis (CF), Fabry's disease (FD), steroid 21-hydroxylase deficiency (21-HD) and Duchenne/Becker muscular dystrophy (DMD/BMD). The method does not require special equipment, labeling or standardization for every PCR product, since conditions of heteroduplex formation and enzyme digestion are universal for all products. The results showed that the method is rapid, effective, safe, reliable, and very simple, as the mutations are visualized on agarose or nusieve/agarose gels. The protocol was furthermore evaluated in three DMD patients with the detection of three alterations which after sequencing, were characterized as disease causative mutations. The proposed assay, which was applied for the first time in a variety of monogenic disorders, indicates that point mutation identification is feasible in any conventional molecular lab even for cases, where other techniques have failed.
...
PMID:Screening human genes for small alterations performing an enzymatic cleavage mismatched analysis (ECMA) protocol. 1795 67

Congenital adrenal hyperplasia (CAH) is a common autosomal recessive disorder which causes more than 90% of CAH cases due to defects in the steroid 21-hydroxylase gene (CYP21A2). The frequency of large mutations was determined in 200 ethnic Chinese (i.e., Taiwanese) CAH patients belonging to 200 families with different clinical forms of CYP21A2 deficiency over 10 years of molecular diagnoses. For a large-gene deletion (or conversion) and the CYP21A2 deletion identification, a PCR product covering the TNXB gene and the 5'-end of the CYP21A2 gene with TaqI endonuclease digestion was analyzed by electrophoresis on agarose gels. For CYP21A2 mutational analysis, secondary PCR amplification of the amplification-created restriction site method was applied. From the results of the analysis, we found that large-gene deletions (or conversions) occurred in 7.5% of the alleles including three different types of the chimeric CYP21A1P/CYP21A2 genes and the haplotype of IVS2-12A/C>G in combination with the 707-714del mutation (without the P30L mutation). The CYP21A2 deletion occurred in 2.0% of the alleles which contained three types of the chimeric TNXA/TNXB genes with two novel ones. We concluded that the CYP21A2 deletion in the ethnic Chinese (Taiwanese) patients exhibits a low occurrence, with the haplotype of the IVS2-12A/C>G in combination with the 707-714del mutation (without the P30L mutation) being prevalent among large gene deletions or conversions.
...
PMID:Low frequency of the CYP21A2 deletion in ethnic Chinese (Taiwanese) patients with 21-hydroxylase deficiency. 1803 88

Many human diseases are caused by small alterations in the genes and in the majority of cases sophisticated protocols are required for their detection. In this study we estimated the efficacy of an enzymatic protocol, which using a new mismatch-specific DNA plant endonuclease from celery (CEL family) recognizes and cleaves mismatched alleles between mutant and normal PCR products. The protocol was standardized on a variety of known mutations, in 11 patients with cystic fibrosis (CF), Fabry's disease (FD), steroid 21-hydroxylase deficiency (21-HD), and Duchenne/Becker muscular dystrophy (DMD/BMD). The method does not require special equipment, labeling or standardization for every PCR product, since conditions of heteroduplex formation and enzyme digestion are universal for all products. The results showed that the method is rapid, effective, safe, reliable, and very simple, as the mutations are visualized on agarose or nusieve/agarose gels. The protocol was furthermore evaluated in three DMD patients with the detection of three alterations, which after sequencing, were characterized as disease causative mutations. The proposed assay, which was applied for the first time in a variety of monogenic disorders, indicates that point mutation identification is feasible in any conventional molecular lab even for cases where other techniques have failed.
...
PMID:Screening human genes for small alterations performing an enzymatic cleavage mismatched analysis (ECMA) protocol. 2393 97


1

---