Biomedical subjects

E Grill

Publications and source records attributed to E Grill.

6 recordsLinked to original sources

PubMed1991

Reactivation of metal-requiring apoenzymes by phytochelatin-metal complexes.

The enzymatically inactive, metal-requiring apoforms of diamino oxidase and of carbonic anhydrase were reactivated by copper-and zinc-phytochelatin complexes, respectively. The level and the rate of reactivation effected by metal complexes consisting of poly(gamma-glutamylcysteinyl)glycine as well as by the respective free metal ion were compared. An efficient transfer of zinc and copper from phytochelatin-complexes to apoenzymes was observed in vitro.

Amine Oxidase (Copper-Containing)↗

PubMed1991

Construction and characterization of a yeast artificial chromosome library of Arabidopsis which is suitable for chromosome walking.

A yeast artificial chromosome (YAC) genomic library of Arabidopsis thaliana was constructed in a derivative of the vector pYAC4 which was modified to facilitate the production of end-specific probes for chromosome walking. Experiments in which a subset of 2300 clones from the library were probed with 30 restriction fragment length polymorphism (RFLP) markers indicated that, on the average, the entire genome is represented once in each 800 YAC clones. Thus, the complete library of more than 20,000 YACs is expected to contain most or all of the Arabidopsis genome with a high probability. The YAC clones examined in the sample had an average insert size of approximately 150 kb +/- 10 and represented more than 5% of the Arabidopsis genome. Based on the properties of the library and the currently available RFLP maps for Arabidopsis, only one or two steps from flanking RFLPs should be sufficient to isolate an average gene in Arabidopsis by chromosome walking with the YACs. In order to facilitate chromosome walking, a method for the production of hybridization probes from the ends of the inserts was employed that is based on a combination of pre-amplification of the vector/insert junction sequences via the polymerase chain reaction and specific transcription from T3 or T7 RNA polymerase promoters flanking the cloning site in the YAC vector.

Base Sequence↗

PubMed1991

Phytochelatins.

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Amino Acid Sequence↗

PubMed1991

Development of a system for efficient chromosome walking in Arabidopsis.

The small genome size of Arabidopsis and the low level of repetitive DNA sequences make this crucifer an attractive system for chromosome walking to isolate genes. Mapping of a mutant locus relative to restriction fragment length polymorphism (RFLP) markers provides the first step towards isolating the corresponding gene. The RFLP marker closest to the target gene serves as a starting point. The distance between gene and marker is generally in the range of 50-200 kb. In order to facilitate chromosome walking of this magnitude, we constructed a yeast artificial chromosome (YAC) library of Arabidopsis. Large fragments of Arabidopsis DNA were cloned into a YAC vector and transformed into yeast. The library contains more than 10 equivalents of the Arabidopsis genome. YACs containing sequences of RFLP markers of Arabidopsis revealed an average insert size of 150 kb. Thus, 1-3 (contiguous) YACs should be sufficient to clone genes from Arabidopsis by chromosome walking. In order to use the system to isolate genes involved in the signal transduction of abscisic acid, we fine-mapped the mutant loci abi-1 and abi-2, which confer abscisic acid insensitivity, relative to RFLPs and isolated the corresponding YACs.

Abscisic Acid↗

PubMed1987

Phytochelatins, the heavy metal binding peptides of plants: characterization and sequence determination.

The isolation of a homologous series of five novel heavy metal complexing peptides from plant cell suspension cultures is reported. Their structures were established as (gamma-Glu-Cys)n Gly, n = 3-7. These peptides appear upon induction with heavy metals and represent the principal metal binding activity in induced cells. The name phytochelatin (PC) is proposed for this new class of natural products in higher plants. The hitherto postulated metallothioneins do not exist in plants but their function of chelating heavy metals via metal-thiolate coordination is fulfilled by the simpler phytochelatins. They are not primary gene products because of their repetition of gamma-glutamyl linkages.

Amino Acid Sequence↗

PubMed1987

Thermostability and superhelicity of plasmid DNA in Bacillus stearothermophilus.

The thermostability of the staphylococcal plasmids pC194 and pUB110 and their antibiotic-resistance determinants was examined upon transfer to Bacillus stearothermophilus CU21. Plasmid pGS13, a pUB110 derivative carrying the chloramphenicol acetyltransferase (CAT) gene of pC194, could be maintained up to the maximum growth temperature (68 degrees C) by selection for chloramphenicol resistance. In the absence of selective pressure, pGS13 was lost at temperatures above 60 degrees C. Segregational instability of pGS13 was accompanied by a progressive loss of negative superhelicity at elevated temperatures. Thermostable mutants of pGS13 were isolated by screening for expression of the antibiotic-resistance determinants after growth under non-selective conditions. These mutants were found to contain an insertion of a 1.7 kb DNA sequence derived from the cryptic B. stearothermophilus plasmid pBS02. Increased thermostability correlated with preservation of plasmid superhelicity at elevated temperatures.

DNA Restriction Enzymes↗