Pseudoachondroplasia due to the substitution of the highly conserved Asp482 by Gly in the seventh calmodulin-like repeat of cartilage oligomeric matrix protein.
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Achondroplasia↗
Biomedical subjects
S Susic
Publications and source records attributed to S Susic.
Multiple epiphyseal dysplasia and pseudoachondroplasia due to novel mutations in the calmodulin-like repeats of cartilage oligomeric matrix protein.
A child with a mild form of pseudoachondroplasia was heterozygous for a deletion of 12 nucleotides from exon 10 of the cartilage oligomeric matrix protein (COMP) gene. It resulted in the deletion of valine 513 to lysine 516 from the eighth calmodulin-like repeat of COMP monomers. A child with the Fairbank's type of multiple epiphyseal dysplasia was also heterozygous for a COMP mutation. It substituted cysteine 371 by serine in the fourth calmodulin-like repeat. Both mutations were likely to alter the conformation and calcium binding of the mutant COMP protein chains. These findings support the proposal that deletions and insertions within the calmodulin-like domain produce pseudoachondroplasia, while amino acid substitutions with this domain may produce either pseudoachondroplasia or multiple epiphyseal dysplasia.
Achondroplasia↗
Human DNA helicase II: a novel DNA unwinding enzyme identified as the Ku autoantigen.
Human DNA helicase II (HDH II) is a novel ATP-dependent DNA unwinding enzyme, purified to apparent homogeneity from HeLa cells, which (i) unwinds exclusively DNA duplexes, (ii) prefers partially unwound substrates and (iii) proceeds in the 3' to 5' direction on the bound strand. HDH II is a heterodimer of 72 and 87 kDa polypeptides. It shows single-stranded DNA-dependent ATPase activity, as well as double-stranded DNA binding capacity. All these activities comigrate in gel filtration and glycerol gradients, giving a sedimentation coefficient of 7.4S and a Stokes radius of approximately 46 A, corresponding to a native molecular weight of 158 kDa. The antibodies raised in rabbit against either polypeptide can remove from the solution all the activities of HDH II. Photoaffinity labelling with [alpha-32P]ATP labelled both polypeptides. Microsequencing of the separate polypeptides of HDH II and cross-reaction with specific antibodies showed that this enzyme is identical to Ku, an autoantigen recognized by the sera of scleroderma and lupus erythematosus patients, which binds specifically to duplex DNA ends and is regulator of a DNA-dependent protein kinase. Recombinant HDH II/Ku protein expressed in and purified from Escherichia coli cells showed DNA binding and helicase activities indistinguishable from those of the isolated protein. The exclusively nuclear location of HDH II/Ku antigen, its highly specific affinity for double-stranded DNA, its abundance and its newly demonstrated ability to unwind exclusively DNA duplexes, point to an additional, if still unclear, role for this molecule in DNA metabolism.
Adenosine Triphosphatases↗
Physiological concentration of magnesium ions induces a strong macroscopic curvature in GGGCCC-containing DNA.
The bending propensity of non-A/T DNA sequence elements is well known, but helical phasing/gel mobility experiments fail to reveal an intensive macroscopic curvature if A/T tracts are not present in the sequence. Recent X-ray data prove on the other hand that a GGCC element is intrinsically curved toward the major groove, which seemingly contradicts the fact that macroscopic curvature at GGGCCC elements is hardly detectable with a conventional gel mobility assay. Here we show that GGGCCC containing DNA, with no A/T tracts in the sequence context, has a detectable, strong gel mobility anomaly only in the presence of divalent ions (10 mM Mg2+ or Ca2+, 1 mM Zn2+). Metal ions increase the gel mobility anomaly in A/T tracts as well, but the effect is substantially stronger for GGGCCC than for the rigid A/T tracts. Our data suggest that metal ions change the sequence-dependent dynamic features of DNA; on the other hand, there is no evidence of twist-mediated change of the planarity of curvature in the presence of metal ions. The results show that near-physiological concentrations of divalent cations (10 mM MgCl2) have a strong and differential effect on various sequence elements, so that the current picture of sequence-dependent DNA curvature is changed not only in a quantitative, but also in a qualitative sense.
Base Composition↗
Evidence for opposite groove-directed curvature of GGGCCC and AAAAA sequence elements.
The repetitive sequence (AGGGCCCTAGAGGGGCCC-TAG)n was previously shown to be curved by gel mobility assays. Here we show, using hydroxy radical/DNase I digestion and differential helical phasing experiments that the curvature is directed towards the major groove and is located in the GGGCCC, but not the CTAGAG segments. The effect of the GC step in the context of the GGGCCC motif is apparently about as large as that of AA/TT, i.e. enough to cancel the macroscopic curvature of helically phased A-tracts. These data are in agreement with positive roll-like curvature of the GCC/GGC motif, predicted from nucleosome packing data and the 3D structure of the GGGGCCCC octamer, but they are not in agreement with the dinucleotide-based roll angle values predicted for AG/CT, TA, GG/CC and GC steps. Our results thus indicate the importance of interactions beyond the dinucleotide steps in predictive models of DNA curvature.
Base Sequence↗
Sequence-specific spin labeling of DNA.
Several DNA fragments deriving from plasmid pBR322 were used to determine the modification sites caused by the reaction with alkylating spin-labeling probes. At a high spin-label concentration, all guanines became alkylated, causing the cleavage of the phosphodiester bonds upon the treatment with piperidine. The lengths of the breakage products of 5'-end labeled DNA treated with spin labels were compared with the length of DNA scission products generated by Maxam-Gilbert procedure for DNA sequence analysis. The distribution of the guanine modifications is dependent on the amount of the reagent used for the alkylation and the ionic conditions of the reaction. The frequency of alkylation by spin labels was greatly enhanced within continuous runs of guanines in DNA. The stabilization of the DNA structure by magnesium or spermine directs the spin-label binding specifically to the most exposed region of DNA fragment containing GGTGG sequence. The sequence-dependent interaction of spin labels with DNA enables the development of the method for the selective spin labeling of DNA molecule.
Alkylation↗