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PubMed2001

Structural and functional analysis of a new desmin variant causing desmin-related myopathy.

Desmin-related myopathy is a familial or sporadic disease characterized by skeletal muscle weakness and cardiomyopathy as well as the presence of intracytoplasmic aggregates of desmin-reactive material in the muscle cells. Previously, two kinds of deletions and eight missense mutations have been identified in the desmin gene and proven to be responsible for the disorder. The present study was conducted to determine structural and functional defects in a pathogenic desmin variant that caused a disabling disorder in an isolated case presenting with distal and proximal limb muscle weakness and cardiomyopathy. We identified a novel heterozygous Q389P desmin mutation located at the C-terminal part of the rod domain as the causative mutation in this case. Transfection of desmin cDNA containing the patient's mutation into C2.7, MCF7, and SW13 cells demonstrated that the Q389P mutant is incapable of constructing a functional intermediate filament network and has a dominant negative effect on filament formation. We conclude that Q389P mutation is the molecular event leading to the development of desmin-related myopathy.

Adult↗
PubMed2005

Diabetes-associated mutations in human insulin: crystal structure and photo-cross-linking studies of a-chain variant insulin Wakayama.

Naturally occurring mutations in insulin associated with diabetes mellitus identify critical determinants of its biological activity. Here, we describe the crystal structure of insulin Wakayama, a clinical variant in which a conserved valine in the A chain (residue A3) is substituted by leucine. The substitution occurs within a crevice adjoining the classical receptor-binding surface and impairs receptor binding by 500-fold, an unusually severe decrement among mutant insulins. To resolve whether such decreased activity is directly or indirectly mediated by the variant side chain, we have determined the crystal structure of Leu(A3)-insulin and investigated the photo-cross-linking properties of an A3 analogue containing p-azidophenylalanine. The structure, characterized in a novel crystal form as an R(6) zinc hexamer at 2.3 A resolution, is essentially identical to that of the wild-type R(6) hexamer. The variant side chain remains buried in a nativelike crevice with small adjustments in surrounding side chains. The corresponding photoactivatable analogue, although of low affinity, exhibits efficient cross-linking to the insulin receptor. The site of photo-cross-linking lies within a 14 kDa C-terminal domain of the alpha-subunit. This domain, unrelated in sequence to the major insulin-binding region in the N-terminal L1 beta-helix, is also contacted by photoactivatable probes at positions A8 and B25. Packing of Val(A3) at this interface may require a conformational change in the B chain to expose the A3-related crevice. The structure of insulin Wakayama thus evokes the reasoning of Sherlock Holmes in "the curious incident of the dog in the night": the apparent absence of structural perturbations (like the dog that did not bark) provides a critical clue to the function of a hidden receptor-binding surface.

Amino Acid Sequence↗
PubMed2005

Molecular models of NS3 protease variants of the Hepatitis C virus.

BACKGROUND: Hepatitis C virus (HCV) currently infects approximately three percent of the world population. In view of the lack of vaccines against HCV, there is an urgent need for an efficient treatment of the disease by an effective antiviral drug. Rational drug design has not been the primary way for discovering major therapeutics. Nevertheless, there are reports of success in the development of inhibitor using a structure-based approach. One of the possible targets for drug development against HCV is the NS3 protease variants. Based on the three-dimensional structure of these variants we expect to identify new NS3 protease inhibitors. In order to speed up the modeling process all NS3 protease variant models were generated in a Beowulf cluster. The potential of the structural bioinformatics for development of new antiviral drugs is discussed. RESULTS: The atomic coordinates of crystallographic structure 1CU1 and 1DY9 were used as starting model for modeling of the NS3 protease variant structures. The NS3 protease variant structures are composed of six subdomains, which occur in sequence along the polypeptide chain. The protease domain exhibits the dual beta-barrel fold that is common among members of the chymotrypsin serine protease family. The helicase domain contains two structurally related beta-alpha-beta subdomains and a third subdomain of seven helices and three short beta strands. The latter domain is usually referred to as the helicase alpha-helical subdomain. The rmsd value of bond lengths and bond angles, the average G-factor and Verify 3D values are presented for NS3 protease variant structures. CONCLUSIONS: This project increases the certainty that homology modeling is an useful tool in structural biology and that it can be very valuable in annotating genome sequence information and contributing to structural and functional genomics from virus. The structural models will be used to guide future efforts in the structure-based drug design of a new generation of NS3 protease variants inhibitors. All models in the database are publicly accessible via our interactive website, providing us with large amount of structural models for use in protein-ligand docking analysis.

Amino Acid Sequence↗
PubMed1987

A histopathologic study of 116 ameloblastomas with special reference to the desmoplastic variant.

The histopathologic features of 116 ameloblastomas were reviewed with special reference to the incidence of the recently described desmoplastic variant. The series included 110 intraosseous lesions and 6 extraosseous ameloblastomas. Ninety-six (88%) of the intraosseous tumors were found in the mandible, and 61% involved the molar-ascending ramus area. Fourteen examples of the desmoplastic variant were identified, representing 13% of the intraosseous tumors. The lesions showed distinctive histologic features characterized by an extensive collagenized stroma containing small islands of tumor epithelium with scant tendency to form cystic structures. This histologic variant has a marked tendency to involve the anterior portions of the jaws with 7 of the 14 examples being located in the maxilla. These desmoplastic lesions also showed unusual radiographic findings, often more suggestive of a benign fibro-osseous lesion than of ameloblastoma.

Adolescent↗
PubMedDate not supplied

[Analysis of genome determinants of virulence and setting-up of a library of full-size genome copies of the attenuated virus strain of the porcine reproductive and respiratory syndrome virus North American type].

Primary genome structures of 3 variants of the NADC-8 North American virulent strain of porcine reproductive and respiratory syndrome virus (PRRSV) were compared for the purpose of detecting any potential genetic virulence determinants of genus Arterivirus. Apart from the virulent variant, we also investigated the attenuated variant, obtained after 251 passages in cell culture, and the intermediate variant isolated from a pig after a partial reversion of the attenuated virus. The attenuated variant genome acquired a 3-nucleotide deletion and 50 mutations versus its virulent precursor. A comparison of the attenuated and intermediary virus variants denoted 8 nucleotide mutations entailing substitutions of 6 amino acids in 3 open reading frames (ORF1a, ORF1b and ORF6). A 32-clone library was constructed in the pACYC177 plasmid vector, which comprised full-size copies of the genome of the NADC-8 attenuated variant strain (251), virus PPCC, for the purpose of experimentally verifying the functional role of the obtained mutations. Full-size analogues ((+)-chain of RNA) of the viral genome, comprising the CAP-structures and polyadenylated ones were obtained in vitro on the basis of the cloned DNA. Seven of the 8 analyzed clones of the viral genome were infected and their insertion into the MARC-145 cell resulted in obtaining of infectious PRRSVs. Four of the constructed recombinant viruses had delayed growth parameters, and 3 of them were similar to the parental strain. The described technology (inverse genetics) would make it possible to introduce changes into the viral genome in applied and fundamental research of Arteriviruses.

Amino Acid Substitution↗
PubMed1985

IEF patterns of HLA-B13 antigens from Orientals and Caucasians.

HLA-B13 antigens were isolated from metabolically labeled cell extracts from Caucasian and Oriental donors by means of an HLA-B13-specific monoclonal antibody, SY1. Ethnic differences in B13 molecules were identified by one-dimensional isoelectric focusing in which the pI of desialated Oriental B13 molecules was found to be higher than that of Caucasians. An additional Caucasian variant pattern was detected by peptide mapping using limited proteolysis in sodium dodecyl sulfate analyzed by polyacrylamide gel electrophoresis. Dual allotypic determinants for B13 molecules were recognized by two HLA-B13-specific monoclonal antibodies, SY1 and Tu110, as determined by their sensitivity to complement-dependent cell lysis. Whereas the SY1 target epitope was shared by both ethnic B13 molecules, the two ethnic B13 molecules carried different Tu110 target structures. The Caucasian variant molecules appear to carry altered allotypic determinants which are recognized by both SY1 and Tu110 antibodies. This study suggests that the HLA-B13 private structure may comprise two epitopes recognized by SY1 and Tü110 antibodies, respectively, whose binding sites overlap. Present data also suggest that the private determinant was already present when the two racial groups diverged, and thus the mutations which gave rise to the variants may be of relatively recent origin.

Antibodies, Monoclonal↗
PubMed1990

Structural characterization of idiotopes by using antibody variants generated by site-directed mutagenesis.

Four anti-idiotopic mAB, 107, MB, AI, and AD8, react with mouse hybridoma protein 36-65 specific for the hapten p-azophenylarsonate. The four antiidiotypic antibodies do not react with hybridoma protein 36-71, a somatically mutated variant of 36-65 whose H and L chain V region sequence differs at 19 amino acid positions. To determine which regions of 36-65 are important for the interaction with each of the four anti-idiotypic antibodies, variants of 36-65 containing one or more of the 36-71 substitutions were generated by oligonucleotide-directed mutagenesis of the rearranged 36-65 H chain V region gene, followed by expression of mutant proteins containing either the 36-65 or the 36-71 L chain in transfected hybridoma cells. Idiotypic characterization of the mutant proteins showed that reactivity correlates with the 36-65 H chain, but some contributions from the 36-65 L chain come into play. In the 36-65 H chain V region, idiotopes were mapped to the first and third complementarity-determining regions for anti-idiotypic antibodies 107, MB, and AI, and to all three complementarity-determining regions for anti-idiotypic antibody AD8. The binding of all four anti-idiotypic antibodies to hybridoma protein 36-65 was hapten inhibitable. However, a comparison between the effect of individual 36-71 substitutions on idiotope expression and their effect on Ag-binding affinity suggests that none of the four anti-idiotypic antibodies bodies mimics the structure of Ag.

Amino Acid Sequence↗
PubMed1994

Structural and functional characterization of two allelic variants of human cystatin D sharing a characteristic inhibition spectrum against mammalian cysteine proteinases.

Human cystatin D is a novel member of the cystatin superfamily of cysteine proteinase inhibitors present in saliva and tears. Two alleles of the cystatin D gene (CST5), encoding protein variants with either Cys or Arg as residue 26 in their 122-residue polypeptide chains, are present in the population. Expression of the two alleles was investigated by immunochemical analyses of the secreted cystatin D in saliva from individuals homozygous for each of the two alleles, with results demonstrating that both are expressed at similar levels. The inhibitory characteristics of the two cystatin D variants were studied, by determination of dissociation equilibrium constants (Ki) for their complexes with papain and with the mammalian cysteine proteinases, cathepsins B, H, L, and S. The results demonstrate that 1) cystatin D has a characteristic inhibition profile since it does not inhibit cathepsin B (Ki > 1 microM), and when compared to cystatin C and all other known cystatins it is a much poorer inhibitor of cathepsin L (mean Ki 25 nM) but binds cathepsin H and S relatively tightly (mean Ki values of 8.5 and 0.24 nM, respectively); and 2) the inhibitory activities of the two cystatin D variants are not significantly different, demonstrating that the presence of an extra cysteine residue in the cystatin D molecule affects neither the stability nor the functional activity of the inhibitor, thus explaining the widespread distribution of the Cys26-cystatin D encoding allele in the population. The inhibitory properties displayed by cystatin D suggest that it has a function in saliva as inhibitor of either endogenous or exogenous enzymes with cathepsin S- or H-like properties.

Alleles↗
PubMed1994

Identification of two point mutations in the gene coding luteinizing hormone (LH) beta-subunit, associated with immunologically anomalous LH variants.

To analyze the structure of LH in three patients with immunologically anomalous LH, the whole coding region of the LH beta-subunit gene was examined. These patients were infertile, and their serum LH levels could not be measured with an immunoassay kit. Immunoblotting of the LH beta-subunit showed no marked changes in the molecular size of LH beta. Genomic DNA was extracted from peripheral lymphocytes of the patients and normal controls, and LH beta genes were amplified by the polymerase chain reaction technique, using primer pairs that are capable of specifically amplifying only the LH beta gene without interference by the CG beta genes. No deletions were observed in the coding regions of the LH beta gene of the patients. Nucleotide sequencing revealed two nucleotide substitutions in the LH beta gene of the patients, which cause amino acid replacements from Trp8 (TGG) to Arg8 (CGG) and Ile15 (ATC) to Thr15 (ACC). Restriction fragment length polymorphism analysis in three families indicated that the affected probands were homozygous, and their family members were heterozygous, except for their husbands. The heterozygotes showed reduced detectability with the LH immunoassay kit. These results suggest that these amino acid replacements are responsible for this immunologically anomalous variant.

Adult↗
PubMed1993

Single-strand conformation polymorphism (SSCP) analysis of the molecular pathology of hemophilia B.

In the present study, we report the application of polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) analysis to the screening of seven functionally important factor IX gene (FIX) regions (total length 2.66 kb) in 9 unrelated haemophilia B patients of Portuguese or African origin. In eight of the patients an altered migration pattern of single-stranded DNA was observed. Direct sequencing of the relevant DNA fragments unveiled the following sequence alterations: two novel mutations, namely FIXBarcelos Thr-380-Pro and FIXLousada 9bp insertion at position 31,309 or 31,318; five mutations previously reported in other ethnic groups (FIXPorto Arg-145-His, FIXLuanda Gly-207-Arg, FIXPenafiel Arg-248-Gln, FIXSesimbra Arg-333-Gln, FIXCascais Arg-333-Stop); and a normal variant, G-->T transvertion at position 6,596 in intron 2. We propose hypothetical models for the generation of the 9 bp duplication (FIXLousada). We have performed molecular modeling studies in order to predict the structure of the variant FIX molecules.

Base Sequence↗
PubMed2006

Visualization, reconstruction, and integration of neuronal structures in digital brain atlases.

Brain atlases are used in neuroanatomy to define the spatial layout of neuronal structures. Their digital variant can serve as a database and common reference frame for integrating data from different biological experiments. This article presents an overview of methods for three-dimensional visualization of neuroanatomical image data, reconstructing neuronal structures from image data, creating digital brain atlases, and registering data in an atlas. This enables analysis of spatial relations between individual structures imaged in different experiments as well as between these structures and the atlas.

Animals↗
PubMed1993

X-ray structures of the antigen-binding domains from three variants of humanized anti-p185HER2 antibody 4D5 and comparison with molecular modeling.

The X-ray structures of 1 Fv and 2 Fab humanized anti-p185HER2 antibody fragments (IgG1-kappa) have been determined at a resolution between 2.7 A and 2.2 A. The antibodies are three different versions of a human antibody framework onto which the antigen recognition loops from a murine antibody (4D5) have been grafted. The sequences of the three versions differ in the framework region at positions L55, H78 and H102. The version 8 Fv fragment crystallizes in space group P2(1) with cell parameters a = 37.6 A, b = 63.4 A, c = 90.2 A, beta = 98.2 degrees, with two molecules per asymmetric unit, and has been refined against data 10.0 A-2.2 A to an R-factor of 18.3%. Versions 4 and 7 Fabs crystallize in space group P1 with cell parameters a = 39.2 A, b = 80.2 A, c = 86.1 A, alpha = 113.1 degrees, beta = 92.7 A, gamma = 102.6 A and two molecules per asymmetric unit. Version 4 has been refined against data 10.0 A-2.5 A resolution to an R-factor of 17.9%. Version 7 has been refined against data 10 A-2.7 A to an R-factor of 17.1%. The X-ray structures have been used to assess the accuracy of structural predictions made via molecular modeling, and they confirm the structural role of certain framework residues and the conformations of five of six complementarity determining regions (CDRS). The average deviation of the model from the X-ray structures is within the range observed among the X-ray structures for 81% of the C alpha atoms. Of the hydrogen bonds common to the X-ray structures, 94% of the main-chain-main-chain and 79% of the main-chain-side-chain ones were predicted by the model. The side-chain conformation was predicted correctly for 79% of the buried residues. The third CDR in the heavy chain is variable, differing by up to 8 A between molecules within an asymmetric unit. The structural relationship between variable domains of light and heavy chains is not significantly altered by the absence of constant domains in the Fv molecule. The antigen-binding potential of an unusual light chain sequence has been confirmed. The arginine at position 66 interacts with the first light chain CDR, but in a fashion somewhat different than predicted. A substitution of a leucine for an alanine side-chain directed between the beta-sheets has only relatively small and local effects.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed2026

First nationwide full-genome characterisation of human-derived Andes virus in Chile: a retrospective genomic epidemiology study.

BACKGROUND: Andes virus (ANDV) is the only hantavirus known to transmit between humans and causes hantavirus cardiopulmonary syndrome in Chile and Argentina. In Chile, ANDV genomic diversity remains incompletely characterised. This study aimed to characterise the genetic diversity, geographical structure, and molecular signatures of ANDV using human clinical samples collected over a 13-year period (2011-24). METHODS: We conducted a retrospective genomic epidemiology study of ANDV infections in Chile. Clinical samples from patients with confirmed ANDV, collected between March 9, 2011, and June 27, 2024, were analysed and sequenced. Clinical and epidemiological data were obtained from diagnostic laboratories and surveillance programmes. Consensus sequences for the S, M, and L segments were generated, and genetic clustering and divergence were assessed using phylogenetic inference and variant calling. FINDINGS: We analysed clinical samples from 58 infected individuals and identified two major genomic variants of ANDV with distinct geographical distributions, defined by regionally structured patterns of nucleotide and amino acid substitutions across the S, M, and L segments: ANDV Chi-North (central Chile) and ANDV-South (southern Chile). No consistent clustering by clinical severity was observed, and no recurrent non-synonymous substitutions were uniquely associated with severe disease. Substitutions previously associated with person-to-person transmission in outbreaks in Argentina were not consistently observed in Chilean sequences, including in four person-to-person transmission cases. Although some substitutions described in ANDV-like viruses were present in the Chi-North lineage, this lineage remained phylogenetically distinct and geographically restricted to central Chile. INTERPRETATION: To our knowledge, this study provides the first nationwide genomic characterisation of human-derived ANDV in Chile. The identification of geographically structured variants indicates that ANDV diversity in Chile is driven by regional diversification rather than clinical outcome. The absence of consistent amino acid signatures associated with disease severity or person-to-person transmission suggests that these phenotypes are unlikely to be explained by viral genetic variation alone. These findings refine current understanding of ANDV evolution and highlight the need for continued integrated genomic surveillance in endemic regions. FUNDING: Agencia Nacional de Investigación y Desarrollo de Chile and National Institutes of Health.

Humans↗
PubMedDate not supplied

[Posterior hyaloid membrane as structural base of growth of neovascular tissue in proliferative diabetic retinopathy].

Morphologic study of 38 specimens of epiretinal membranes obtained during transciliary vitrectomy in 26 patients with type I diabetes mellitus and 12 with type II diabetes with proliferative diabetic retinopathy of different stages was carried out. In proliferative diabetic retinopathy, new vessels grow mainly along the posterior (retinal) surface of the posterior hyaloid membrane. Sometimes, if there are defects in the posterior hyaloid membrane, proliferative tissue stratifies it and appears on the anterior (hyaloid) surface. We observed no growth of new vessels outside the posterior hyaloid membrane. The impossibility of clinical and mechanical separation of the posterior hyaloid membrane and tumor tissue permits us to regard the stages of proliferative process in diabetic retinopathy as stages in the alteration of the posterior hyaloid membrane proper. Five variants in the structure of proliferative tissue can be distinguished in diabetic retinopathy; we consider that these variants are stages in its development: 1) glial--with predominance of cell-free or hypocellular glial tissue; 2) glial vascular--with growth of thin-wall vessels into glial tissue; 3) glial vascular fibrous--with growth of porous fibrovascular membrane in glial tissue; 4) fibrovascular--with predominance of fibrovascular tissue; and 5) fibrous (cicatricial) hypocellular compact connective tissue with just few vessels or none at all. Reduction of new vessels, involution of proliferative tissue, and development of traction detachment of the retina are associated with the beginning of fibrous proliferation.

Diabetes Mellitus, Type 1↗
PubMed1994

Localization of blood-group-related linear poly-N-acetyllactosamine structure in different human tissues by Griffonia simplicifolia agglutinin-II staining following endo-beta-galactosidase digestion.

Endo-beta-galactosidase from Escherichia freundii cleaves polylactosaminyl structures as follows: R-GlcNAc beta I-3Gal beta I-4GlcNac beta I-R' + H2O-->R-GlcNAc beta I-3Gal + GlcNAc beta I-R'. By staining with Griffonia simplicifolia agglutinin-II following the enzyme digestion, the distribution of R-GlcNAc beta I-3Gal beta I-4GlcNAc can be demonstrated in tissue sections. This carbohydrate chain is one of the backbone structures carrying the blood-group-related antigens and, thus, localization of this structure may provide detailed information about the distribution of variants with different backbone structures. Various formalin-fixed, paraffin-embedded tissue sections were stained by Griffonia simplicifolia agglutinin-II with or without prior enzyme digestion and the reactivity of the agglutinin imparted by enzyme digestion was studied in the following tissues and cells: pancreatic acinar cells, gastric surface mucosae, duct cells and mucous cells of salivary glands and tracheal glands, surface epithelium of trachea, goblet cells of large intestine, columnar epithelium of uterine cervical glands, distal and collecting tubules of kidney, certain cells of anterior lobe and colloid of middle lobe of pituitary glands, epithelial reticular cells and Hassall's corpuscles of thymus and Kupffer cells of liver. In gastric surface mucosae, the reactivity of the agglutinin appeared in non-secretor individuals but not in the secretor individuals, and in mucous cells of salivary and tracheal glands the reactivity appeared in Le(a- b-) non-secretor individuals but not in Le(a + b-) non-secretor or secretor individuals.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed2024

The characterization of new de novo CACNA1G variants affecting the intracellular gate of Cav3.1 channel broadens the spectrum of neurodevelopmental phenotypes in SCA42ND.

PURPOSE: Missense de novo variants in CACNA1G, which encodes the Cav3.1 T-type calcium channel, have been associated with a severe, early-onset form of cerebellar disorder with neurodevelopmental deficits (SCA42ND). We explored a large series of pediatric cases carrying heterozygous variants in CACNA1G to further characterize genotype-phenotype correlations in SCA42ND. METHODS: We describe 19 patients with congenital CACNA1G-variants, including 6 new heterozygotes of the recurrent SCA42ND variants, p.(Ala961Thr) and p.(Met1531Val), and 8 unreported variants, including 7 missense variants, mainly de novo. We carried out genetic and structural analyses of all variants. Patch-clamp recordings were performed to measure their channel activity. RESULTS: We provide a consolidated clinical description for the patients carrying p.(Ala961Thr) and p.(Met1531Val). The new variants associated with the more severe phenotypes are found in the Cav3.1 channel intracellular gate. Calcium currents of these Cav3.1 variants showed slow inactivation and deactivation kinetics and an increase in window current, supporting a gain of channel activity. On the contrary, the p.(Met197Arg) variant (IS4-S5 loop) resulted in a loss of channel activity. CONCLUSION: This detailed description of several de novo missense pathogenic variants in CACNA1G, including 13 previously reported cases, supports a clinical spectrum of congenital CACNA1G syndrome beyond spinocerebellar ataxia.

Humans↗
PubMed1980

Proteus mirabilis phages 5006M, 5006M HFT k and 5006M HFT ak: physical comparison of genome characteristics.

The genomes of proteus mirabilis phages 5006M, kanamycin resistance transducing variant 5006M HFT k and kanamycin-ampicillin resistance transducing variant 5006M HFT ak have been compared. Homo- and heteroduplex and partial denaturation mapping analyses were performed. The results confirm a sequential headful packaging mechanism, facilitate mapping of the ampicillin resistance marker, demonstrate a hairpin loop structure in both variants, reveal a common insertion site for 8 X 10(6) mol. wt. non-phage DNA in both variants and implicate a role for the non-inducible cryptic host strain prophage 5006M in the generatin cycle of the variant phages.

Ampicillin↗
PubMed1995

Phase variation of Enterococcus faecalis pAD1 conjugation functions relates to changes in iteron sequence region.

pAD1 (60 kb) is a conjugative, hemolysin/bacteriocin plasmid in Enterococcus faecalis. It confers a mating response to the peptide sex pheromone cAD1 produced by recipient (plasmid-free) cells, leading to highly efficient plasmid transfer in broth matings. Control of the physiological response to cAD1 can been overridden by a reversible phase variation event at frequencies on the order of 10(-4) to 10(-3) per cell per generation (L. T. Pontius and D. B. Clewell, Plasmid 26:172-185, 1991). The variant forms are designated Dryc and Dry+, which reflects the colony morphologies of cells whose conjugation functions are switched on and off, respectively. Here we show that Dryc variants exhibit a structural change in a region between repA and repB that contains two clusters of 8-bp iterons. The change involved a 31- or 32-bp increase in size of this region. In three or four independent variants examined, one of the iteron clusters increased in size from 13 to 17 iterons. When iteron DNA was placed on a multicopy plasmid and introduced into a wild-type pAD1 derivative, the Dryc phenotype was generated. Since traA, a key negative regulator of conjugation, bears several centrally located iteron-like sequences with the same orientation, we speculate that the protein(s) that normally binds iterons (possibly RepA and/or RepB) blocks traA transcription in Dryc variants.

Bacterial Proteins↗