Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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)
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PMID:Exon 7 Ncol restriction site within CYP21B (steroid 21-hydroxylase) is a normal polymorphism. 197 47

Steroid 21-hydroxylase deficiency is the leading cause of impaired cortisol synthesis in congenital adrenal hyperplasia (CAH). We have studied the structure of the CYP21B gene in 30 unrelated CAH patients using the polymerase chain reaction (PCR) to differentiate the active CYP21B gene from its highly related CYP21A pseudogene. The PCR approach obviates the need to distinguish the CYP21A and CYP21B genes by restriction endonuclease digestion and electrophoresis before analysis with labeled probes. Furthermore, direct nucleotide sequence analysis of CYP21B genes is demonstrated on the PCR-amplified DNA. Gene deletion of CYP21B, gene conversion of the entire CYP21B gene to CYP21A, frame shift mutations in exon 3, an intron 2 mutation that causes abnormal RNA splicing, and a mutation leading to a stop codon in exon 8 appear to be the major abnormalities of the CYP21B gene in our patients. These mutations appear to account for 21-hydroxylase deficiency in 22 of 26 of our salt-wasting CAH patients.
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PMID:Direct analysis of CYP21B genes in 21-hydroxylase deficiency using polymerase chain reaction amplification. 232 62

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.
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PMID:Clinical and genetic characterization of nonclassic 21-hydroxylase deficiency. 278 81

Congenital adrenal hyperplasia (CAH) is caused by disorders of the P450c21B gene, which, with the P450c21A pseudogene, lies in the HLA locus on chromosome 6. The near identity of nucleotide sequences and endonuclease cleavage sites in these A and B loci makes genetic analysis of this disease difficult. We used a genomic DNA probe that detects the P450c21 genes (A pseudogene, 3.2 kb; B gene, 3.7 kb in Taq I digests) and the 3' flanking DNA not detected with cDNA probes (A pseudogene, 2.4 kb; B gene, 2.5 kb) to examine Southern blots of genomic DNA from 68 patients and 165 unaffected family members in 57 families with CAH. Of 116 CAH-bearing chromosomes, 114 could be sorted into five easily distinguished haplotypes based on blots of DNA digested with Taq I and Bgl II. Haplotype I (76 of 116, 65.6%) was indistinguishable from normal and therefore bore very small lesions, presumably point mutations. Haplotype II (4 of 116, 3.4%) and haplotype III (8 of 116, 6.9%) had deletions and duplications of the P450c21A pseudogene but had structurally intact P450c21B genes presumably bearing point mutations; point mutation thus was the genetic defect in 88 of 116 chromosomes (75.9%). Haplotypes IV and V lack the 3.7-kb Taq I band normally associated with the P450c21B gene. Haplotype IV (13 of 116, 11.2%) retains all other bands, indicating that the P450c21B gene has undergone a gene conversion event, so that it is now also associated with a 3.2-kb band. Haplotype V (13 of 116, 11.2%) lacks the 2.4-kb Taq I fragment and the 12-kb Bgl II fragments normally associated with the P450c21A pseudogene, as well as lacking the 3.7-kb Taq I fragment, indicating deletion of approximately 30 kb of DNA, resulting in a single hybrid P450c21A/B gene. Most (114 of 116, 98%) CAH alleles thus can easily be classified with this new probing strategy, eliminating many ambiguities resulting from probing with cDNA.
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PMID:Rearrangements and point mutations of P450c21 genes are distinguished by five restriction endonuclease haplotypes identified by a new probing strategy in 57 families with congenital adrenal hyperplasia. 291 51

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.
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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.
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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.
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PMID:Gene conversion-like events cause steroid 21-hydroxylase deficiency in congenital adrenal hyperplasia. 350 Apr 73