SEARCH · PubMed Health

Results for “insertion, inversion”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 487 records · Page 27Linked to original sources

PubMed2005

Probing the environment of signal-anchor sequences during topogenesis in the endoplasmic reticulum.

Signal sequences for insertion of protein into the mammalian endoplasmic reticulum orient themselves in the translocon on the basis of their flanking charges. It has recently been shown that hydrophobic N-terminal signals initially insert head-on before they invert their orientation to translocate the C-terminus. The rate of inversion is reduced with the increasing hydrophobicity of the signal due to an increased affinity for the initial bound state at the translocon. To probe the environment of the signal while its orientation is determined, different hydrophobic residues were inserted at various positions throughout a uniform oligoleucine signal sequence and the constructs were expressed in transfected COS-7 cells. The resulting topologies revealed a strikingly symmetric position dependence specifically for bulky aromatic amino acids, reflecting the structure of a lipid bilayer. Maximal N-translocation was observed when the guest residues were placed at the N- or C-terminus of the hydrophobic sequence or in the very center, corresponding to the positions of highest expected affinity of the signal sequence as a membrane-spanning helix for the bilayer. The results support the model that during topogenesis in vivo the signal sequence is exposed to the lipid membrane.

Alanine↗

PubMed2002

Use of the transposon Ac as a gene-searching engine in the maize genome.

We show here that, although genes constitute only a small percentage of the maize genome, it is possible to identify them phenotypically as Ac receptor sites. Simple and efficient Ac transposition assays based on the well-studied endosperm markers bz and wx were used to generate a collection of >1300 independent Ac transposants. The majority of transposed Ac elements are linked to either the bz or the wx donor loci on chromosome 9. A few of the insertions produce obvious visible phenotypes, but most of them do not, suggesting that these populations will be more useful for reverse genetics than for forward transposon mutagenesis. An inverse polymerase chain reaction method was adapted for the isolation of DNA adjacent to the transposed Ac elements (tac sites). Most Ac insertions were into unique DNA. By sequencing tac sites and comparing the sequences to existing databases, insertions were identified in a number of putative maize genes. The expression of most of these genes was confirmed by RNA gel blot analysis. We report here the isolation and characterization of the first 46 tac sites from the two insertion libraries.

Computational Biology↗

PubMed1997

Bi-stranded, multisite replication of a base pair between difluorotoluene and adenine: confirmation by 'inverse' sequencing.

BACKGROUND: The nonpolar nucleoside of difluorotoluene (F) was previously found to behave similarly to thymidine in single-site deoxynucleoside triphosphate (dNTP) insertion experiments with the Klenow fragment (KF) of DNA polymerase I. Further study was needed, first to see whether F-A base pairs could be replicated in more than one sequence context; second to investigate whether specific base pair replication occurs in the presence of four dNTPs; and third to confirm the presence of F in a replicated DNA strand. RESULTS: A primer bound to a template strand containing eight F residues was extended by KF using the four natural dNTPs at 20 microM. Similarly, the complement (containing eight adenines) was extended using dATP, dGTP, dCTP and dFTP. Comparison of the new strands to authentic strands using standard and 'inverse' chemical sequencing showed identical composition within +/- 5%. CONCLUSIONS: The results confirm that F in a template strand encodes the insertion of dATP and that adenine in a template encodes the insertion of dFTP with good specificity in at least six different nearest neighbor contexts. The results confirm that analog F behaves similarly to thymidine despite its poor hydrogen-bonding ability.

Adenine↗

PubMed2004

Detailed analysis of the insertion site of the mobile elements R997, pMERPH, R392, R705 and R391 in E. coli K12.

The IncJ group of mobile elements have not been extensively studied until recently, due to the inability to isolate extrachromosomal DNA from IncJ-strains. Sequence analysis of the prototype IncJ element, R391, revealed it to be a mosaic structure, integrated into the prfC gene in E. coli. Using inverse PCR (iPCR), we localised the other available IncJ elements (R392, R705, R997 and pMERPH) site of insertion to a 17-bp sequence, within the 5' end of prfC at 99.31 min on the E. coli chromosome, and confirmed this for R391. Despite disrupting prfC, the IncJ's encode novel promoter and 5' sequences, restoring function of the disrupted prfC. Sequence analysis of the elements ends revealed that they contain integrase genes, which share extensive homologies among the group, despite being isolated from broad geographic locations. The elements excise from the host chromosome by recombination between their attL and attR sites, with subsequent recombination between the attP sites on the circular forms and the attB sites in the host genomes. The attB site is highly conserved and found in many different bacteria, suggesting a possible broad host range.

Amino Acid Motifs↗

PubMed1993

P nucleotides in V(D)J recombination: a fine-structure analysis.

Antigen receptor genes acquire junctional inserts upon assembly from their component, germ line-encoded V, D, and J segments. Inserts are generally of random sequence, but a small number of V-D, D-J, or V-J junctions are exceptional. In such junctions, one or two added base pairs inversely repeat the sequence of the abutting germ line DNA. (For example, a gene segment ending AG might acquire an insert beginning with the residues CT upon joining). It has been proposed that the nonrandom residues, termed "P nucleotides," are a consequence of an obligatory end-modification step in V(D)J recombination. P insertion in normal, unselected V(D)J joining products, however, has not been rigorously established. Here, we use an experimentally manipulable system, isolated from immune selection of any kind, to examine the fine structure of V(D)J junctions formed in wild-type lymphoid cells. Our results, according to statistical tests, show the following, (i) The frequency of P insertion is influenced by the DNA sequence of the joined ends. (ii) P inserts may be longer than two residues in length. (iii) P inserts are associated with coding ends only. Additionally, a systematic survey of published P nucleotide data shows no evidence for variation in P insertion as a function of genetic locus and ontogeny. Together, these analyses establish the generality of the P nucleotide pattern within inserts but do not fully support previous conjectures as to their origin and centrality in the joining reaction.

Animals↗

PubMed1997

Quantitation of chemical exchange rates using pulsed-field-gradient diffusion measurements.

A new approach to the quantitation of chemical exchange rates is presented, and its utility is illustrated with application to the exchange of protein amide protons with bulk water. The approach consists of a selective-inversion exchange HMQC experiment in which a short spin echo diffusion filter has been inserted into the exchange period. In this way; the kinetics of exchange are encoded directly in an apparent diffusion coefficient which is a function of the position of the diffusion filter in the pulse sequence. A detailed theoretical analysis of this experiment indicates that, in addition to the measurement of simple exchange rates, the experiment is capable of measuring the effect of mediated exchange, e.g. the transfer of magnetization from bulk water to an amide site mediated by an internal bound water molecule or a labile protein side-chain proton in fast exchange with bulk water. Experimental results for rapid water/amide exchange in acyl carrier protein are shown to be quantitatively consistent with the exchange rates measured using a selective-inversion exchange experiment.

Acyl Carrier Protein↗

PubMed2001

Conserved amino acid motifs from the novel Piv/MooV family of transposases and site-specific recombinases are required for catalysis of DNA inversion by Piv.

Piv, a site-specific invertase from Moraxella lacunata, exhibits amino acid homology with the transposases of the IS110/IS492 family of insertion elements. The functions of conserved amino acid motifs that define this novel family of both transposases and site-specific recombinases (Piv/MooV family) were examined by mutagenesis of fully conserved amino acids within each motif in Piv. All Piv mutants altered in conserved residues were defective for in vivo inversion of the M. lacunata invertible DNA segment, but competent for in vivo binding to Piv DNA recognition sequences. Although the primary amino acid sequences of the Piv/MooV recombinases do not contain a conserved DDE motif, which defines the retroviral integrase/transposase (IN/Tnps) family, the predicted secondary structural elements of Piv align well with those of the IN/Tnps for which crystal structures have been determined. Molecular modelling of Piv based on these alignments predicts that E59, conserved as either E or D in the Piv/MooV family, forms a catalytic pocket with the conserved D9 and D101 residues. Analysis of Piv E59G confirms a role for E59 in catalysis of inversion. These results suggest that Piv and the related IS110/IS492 transposases mediate DNA recombination by a common mechanism involving a catalytic DED or DDD motif.

Amino Acid Motifs↗

PubMed2005

Sequence determinants of a transmembrane proton channel: an inverse relationship between stability and function.

The driving forces behind the folding processes of integral membrane proteins after insertion into the bilayer, is currently under debate. The M2 protein from the influenza A virus is an ideal system to study lateral association of transmembrane helices. Its proton selective channel is essential for virus functioning and a target of the drug amantadine. A 25 residue transmembrane fragment of M2, M2TM, forms a four-helix bundle in vivo and in various detergents and phospholipid bilayers. Presented here are the energetic consequences for mutations made to the helix/helix interfaces of the M2TM tetramer. Analytical ultracentrifugation has been used to determine the effect of ten single-site mutations, to either alanine or phenylalanine, on the oligomeric state and the free energy of M2TM in the absence and the presence of amantadine. It was expected that many of these mutations would perturb the M2TM stability and tetrameric integrity. Interestingly, none of the mutations destabilize tetramerization. This finding suggests that M2 sacrifices stability to preserve its functions, which require rapid and specific interchange between distinct conformations involved in gating and proton conduction. Mutations might therefore restrict the full range of conformations by stabilizing a given native or non-native conformational state. In order to assess one specific conformation of the tetramer, we measured the binding of amantadine to the resting state of the channel, and examined the overall free energy of assembly of the amantadine bound tetramer. All of the mutations destabilized amantadine binding or were isoenergetic. We also find that large to small residue changes destabilize the amantadine bound tetramer whereas mutations to side-chains of similar volume stabilize this conformation. A structural model of the amantadine bound state of M2TM was generated using a novel protocol that optimizes a structure for an ensemble of neutral and disruptive mutations. The model structure is consistent with the mutational data.

Amantadine↗

PubMed1997

Investigation of an outbreak of multidrug resistant tuberculosis among renal patients using rpo B gene sequencing and IS6110 inverse PCR.

A cluster of cases of tuberculosis among five patients receiving treatment for renal failure was investigated. Insertion sequence (IS6110) fingerprinting and antibiotic resistance profiling of the Mycobacterium tuberculosis isolates from four of the patients (A-D), who had been on the same ward, showed that three of these cases (A-C) were related, but that the fourth (D) was distinct. An isolate from the fifth patient (E), who had been on a separate ward, was indistinguishable from the outbreak strain by IS6110 profile. However, the isolate from patient E and a second isolate from patient A differed from the previous strains in being rifampicin resistant. Sequence analysis of the rpo B genes of the two rifampicin-resistant strains demonstrated the presence of different mutations, showing that they had evolved independently from the same source strain. IS6110 and rpo B gene analyses are invaluable for the accurate investigation of outbreaks of multidrug resistant tuberculosis.

Cross Infection↗

PubMed1974

The manifestation of chromosome rearrangements in unordered asci of Neurospora.

Rapid, effective techniques have been developed for detecting and characterizing chromosome aberrations in Neurospora by visual inspection of ascospores and asci. Rearrangements that are detectable by the presence of deficient, nonblack ascospores in test crosses make up 5 to 10% of survivors after UV doses giving 10-55% survival. Over 135 rearrangements have been diagnosed by classifying unordered asci according to numbers of defective spores. (These include 15 originally identified or analyzed by other workers.) About 100 reciprocal translocations (RT's) have been confirmed and mapped genetically, involving all combinations of the seven chromosomes. Thirty-three other rearrangements generate viable nontandem duplications in meiosis. These consist of insertional translocations (IT's) (15 confirmed), and of rearrangements that involve a chromosome tip (10 translocations and 3 pericentric inversions). No inversion has been found that does not include the centromere. A reciprocal translocation was found within one population in nature. When pairs of RT's that involve the same two chromosome arms were intercrossed, viable duplications were produced if the breakpoints overlapped in such a way that pairing resembled that of insertional translocations (27 combinations).-The rapid analytical technique depends on the following. Deficiency ascospores are usually nonblack (W: "white") and inviable, while nondeficient ascospores, even those that include duplications, are black (B) and viable. Thus RT's typically produce 50% black spores, and IT's 75% black. Asci are shot spontaneously from ripe perithecia, and can be collected in large numbers as groups of eight ascospores representing unordered tetrads, which fall into five classes: 8B:0W; 6B:2W, 4B:4W, 2B:6B, 0B:8W. In isosequential crosses, 90-95% of tetrads are 8:0. When a rearrangement is heterozygous, the frequencies of tetrad classes are diagnostic of the type of rearrangement, and provide information also on the positions of break points. With RT's, 8:0 (alternate centromere segregation) = 0:8 (adjacent-1), 4:4's require interstitial crossing over in a centromere-break point interval, and no 6:2's or 2:6's are expected. With IT's, duplications are viable, 8:0 = 4:4, 6:2's are from interstitial crossing over, 0:8's or 2:6's are rare. Tetrads from RT's that involve a chromosome tip resemble those from IT's, as do tetrads from intercrosses between partially overlapping RT's that involve identical chromosome arms.-Because viable duplications and other aneuploid derivatives regularly occur among the offspring of rearrangements such as insertional translocations, care must be taken in selecting stocks, and original strains should be kept for reference.

Aneuploidy↗

PubMed2006

CSPP and CSPP-L associate with centrosomes and microtubules and differently affect microtubule organization.

We recently described the identification of a centrosome/spindle pole associated protein, CSPP, involved in cell cycle progression. Here we report a CSPP isoform denoted CSPP-L, with a 294 amino acids longer N-terminus and a 51 amino acids insertion located in the coiled-coil mid-domain. Expression analysis indicates an inverse cell cycle dependent regulation. CSPP mRNA expression is highest in G1 whereas CSPP-L expression is highest in G2/M. Ectopic expression of CSPP-L impairs cell cycle progression weaker in G1 than CSPP. Furthermore, normal mitotic phenotypes were observed in CSPP-L but not in CSPP transfectants. CSPP-L relocates from spindle microtubules and poles in metaphase to the mid-spindle in anaphase and concentrates at the mid-body in telophase/cytokinesis. CSPP-L high-expressing mitotic cells were predominantly characterized by lagging chromosomes or monopolar spindles, in contrast to the predominant multipolar spindles observed with CSPP expression. The different effects of CSPP and CSPP-L on microtubule organization in mitosis depend on the coiled-coil mid-domain insertion. The common C-terminal domain is required to repress that activity until mitosis. Notably, this C-terminal domain alone can associate with centrosomes in a microtubule independent manner. Taken together, CSPP and CSPP-L interact with centrosomes and microtubules and can differently affect microtubule organization.

Cell Cycle↗

PubMed2006

Construction of pDESTR, a GATEWAY vector for gene disruption in filamentous fungi.

We have constructed pDESTR, a destination vector of gateway system especially for gene targeting and disruption in filamentous fungi. The vector was constructed by removing the multicloning site of pGEM-T easy vector, and inserting hygromycin phosphotransferase gene construct from pCB1004, and a gateway vector conversion cassette. In order to construct a DNA for gene disruption, only an inverse-polymerase chain reaction (PCR) amplification of the restricted, target sequence is needed. After the amplification with a 5'CACC-tagged primer and an ordinary primer, the DNA fragment will be inserted into pENTR/D-TOPO vector and then transferred into pDESTR through LR-recombination reaction. The resulting vector has the disruption construct, after being digested with the restriction enzyme used for the inverse-PCR. The effectiveness of this vector was assessed in Neurospora crassa. The use of pDESTR will therefore simplify the construction of a targeting vector, where multiple ligation steps are usually needed.

Ascomycota↗

PubMed1981

A complex four-break rearrangement between chromosomes 4 and 13 resulting in a recombinant chromosome 4.

A complex four-break rearrangement between chromosomes 4 and 13 was ascertained in a 10-year-old mentally retarded girl. The rearrangement was inherited from the phenotypically normal mother, who had an inverted insertion of part of the long arm of chromosome 4 into the long arm of 13 and, in addition, a pericentric inversion of the deleted 4. Meiotic crossing-over between the normal and the inverted 4 resulted in a recombinant chromosome 4, which was inherited by the proband, together with the 13/4 insertion. In this way the proband became monosomic for 4q35 leads to qter and trisomic for 4pter leads to 4p15, but she showed only minor physical malformations, as compared with other reports on the trisomy 4p syndrome. The cytogenetic findings are difficult to describe using the ISCN nomenclature.

Chromosome Aberrations↗

PubMed1998

Correction of recurrent nipple inversion with cartilage graft.

Of the many techniques advocated to correct nipples, all demonstrate a failure rate with a recurrence of the inversion. We report a technique to correct the recurrent nipple inversion in a simple and permanent fashion. This technique consists of releasing all scar tissue and any lactiferous ducts deep to the nipple and inserting a cartilage graft (rib or auricular) under the nipple. This cartilage graft serves to fill in any soft tissue defect and prevent recurrent contraction and nipple inversion. Although generally used for correction of recurrent nipple inversion, we have also used this technique successfully as an initial modality in patients with inverted nipples. We have found this technique to be simple, nondeforming, cosmetically acceptable, and permanent in the treatment of both inverted and recurrent inverted nipples.

Adult↗

PubMed1975

Occurrence of insertion sequence (IS) regions on plasmid deoxyribonucleic acid as direct and inverted nucleotide sequence duplications.

Insertion sequence (IS) regions have been identified previously as a cause of strongly polar mutations in Escherichia coli and several bacteriophages. The present experiments indicate that genetically characterized IS regions occur on bacterial plasmid deoxyribonucleic acid (DNA) as both direct and inverted DNA sequence duplications. The DNA insertion which has been shown previously (Sharp et al., 1973) to control expression of tetracycline resistance in the R6-5 plasmid, and which occurs as directly and inversely repeated DNA sequences adjacent to the region believed to contain the tetracycline resistance gene, has been identified as IS3. A second genetically characterized insertion sequence (IS1) has been identified as a direct DNA duplication occurring at both junctions of the resistance transfer factor and R-determinant components of R6-5 and related plasmids. A model is presented for the reversible dissociation of resistance transfer factor and R-determinant components of co-integrate R plasmids at the sites of DNA sequence homology provided by the repeated IS regions.

Base Sequence↗

PubMed2003

[Reconstruction of the anomalous palatoglossal arch causing dysphagia: a case report].

Physical examination of a 20-year-old male patient suffering from dysphagia revealed an abnormally short and medially displaced palatoglossal arch. The mucosa was released by an inverse V incision, and the palatoglossal muscle was found to be located more laterally and cephalad than its usual insertion, attached to the posterior edge of the hard palate. The muscle was transposed medially and caudally to its normal localization on the soft palate, and the palatoglossal arch was reconstructed by closure of the mucosal incision in an inverse Y manner. The patient's complaint disappeared and no complications were observed in the postoperative period.

Adult↗

PubMed2006

Promoter activity of a putative pollen monosaccharide transporter in Petunia hybrida and characterisation of a transposon insertion mutant.

For the growth of the male reproductive cells of plants, the pollen, the presence of sufficient sucrose or monosaccharides is of vital importance. From Petunia hybrida a pollen-specific putative monosaccharide transporter designated PMT1 (for petunia monosaccharide transporter) has been identified previously. The present work provides an in-depth analysis and characterisation of PMT1 in the context of pollen development with the GUS reporter gene and an insertion mutant. The promoter of the pollen-specific putative PMT1 gene has been isolated by inverse PCR and sequenced. Analysis of plants transformed with the promoter-GUS fusion confirmed the specificity of this gene, belonging to the late pollen-specific expressed genes. GUS activity was detected even after 24 h of in vitro pollen germination, at the pollen tube tip. To elucidate the importance of PMT1 for gametophyte development and fertilisation, we isolated a mutant plant containing a transposon insertion in the PMT1 gene by the dTph1 transposon-tagging PCR-based assay. The PMT1 mutant contained a dTph1 insertion in position 1474 bp of the transcribing part of the gene, before the last two transmembrane-spanning domains. Analysis of the progeny of the heterozygous mutant after selfing revealed no alterations in pollen viability and fertility. Mature pollen grains of a plant homozygous for the transposon insertion were able to germinate in vitro in a medium containing sucrose, glucose, or fructose, which indicates that PMT1 is not essential for pollen survival. Several explanations for these results are discussed in the present work.

Amino Acid Sequence↗

PubMedDate not supplied

[Genetic engineering of peptide hormones. II. Possible polymorphism of bovine preprolactin. Molecular cloning data].

The primary structure of an insert from a clone isolated from the bovine pituitary cDNA library by hybridization with prolactin-specific probe has been determined. It was found that the rearrangement of cDNA took place in the process of cloning. The rearrangement includes the inversion of 5'-terminal and the deletion of the central part of cDNA. However from the structure of the insert we were able to deduce the sequences of 5'- and 3'-terminal regions of bovine preprolactin mRNA (257 and 551 bases long). The comparison of these sequences with those published earlier revealed several differences in the primary structure. The most essential of them is the additional triplet coding for alanine in position of -22 of the signal peptide. The heterogeneity of bovine preprolactin mRNA in the region coding for the signal peptide is considered to be a consequence of alternative splicing as it was shown for rat preprolactin mRNA.

Amino Acid Sequence↗