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N Geisler

Publications and source records attributed to N Geisler.

50 records · Page 3Linked to original sources
PubMed1981

Comparison of the proteins of two immunologically distinct intermediate-sized filaments by amino acid sequence analysis: desmin and vimentin.

Although all intermediate-size filaments (10-nm filaments) seem to show similar morphology and share a number of biochemical properties, different cell- and tissue-specific subclasses have been distinguished by immunological experiments and by differences in apparent molecular weights and isoelectric points of the major constituent proteins. In order to understand the degree of possible homology between these proteins, we have begun amino acid sequence analysis of the polypeptides. Here we characterize a large fragment of chicken gizzard and pig stomach desmin as well as the corresponding fragment from porcine eye lens vimentin. The fragments are situated at the carboxyl end and consist of 138-140 amino acid residues--i.e., some 28% of the corresponding polypeptide chains. The results show that the two immunologically distinct porcine proteins are different gene products. They show a related amino acid sequence but differ in 36% of the residues present in the carboxy-terminal region. Thus tissue specificity overrides species divergence. These results are discussed in the light of previous immunological experiments. They lend further support to the hypothesis that intermediate filaments belong to a multigene family, which is expressed in line with certain rules of differentiation during embryogenesis.

Amino Acid Sequence↗
PubMed1980

Purification of smooth-muscle desmin and a protein-chemical comparison of desmins from chicken gizzard and hog stomach.

A fast and convenient procedure for the purification of polymerization-competent smooth-muscle desmin is described. Desmin from chicken gizzard and hog stomach were compared by fingerprint techniques. The two proteins show extensive amino acid sequence homology, although some clear differences in the peptide patterns are indicated. Comparative amino acid sequence analysis of some of the peptides obtained in pure form directly proves this conclusion.

Amino Acid Sequence↗
PubMed1977

Amino-terminal fragments of Escherichia coli lac repressor bind to DNA.

The N-terminal fragments (residues 1-51 and 1-59) obtained by selective tryptic cleavage of native lac repressor retain the ability to bind DNA. These fragments (headpieces) are monomeric and form complexes which resemble those of tetrameric repressor with non-operator DNA. But, they do not show the high specificity of repressor for operator sequences. The DNA binding has been demonstrated by filter-binding assay as well as in solution using absorption, circular dichroism, and fluorescence measurements.

Amino Acid Sequence↗
PubMed1977

Isolation of amino-terminal fragment of lactose repressor necessary for DNA binding.

lac repressor can be dissected by trypsin into a homogenous tetrameric core (accounting for residues 60 to 347), carrying inducer binding activity, and the monomeric amino-terminal peptides ("headpieces") accounting for residues 1 to 59 and 1 to 51, respectively. This restriction of the action of trypsin on lac repressor is obtained in 1 M Tris-HCl (pH 7.5)-30% in glycerol at 25 degrees C since only the peptide bonds at lysine-59 and to a lesser extent after at arginine-51 are cleaved under these conditions. The headpieces can be purified by gel filtration. They have ordered secondary structure as revealed by circular dichroism studies. The monomeric headpieces show the relatively weak binding to nonoperator DNA but not the highly specific and strong binding to operator DNA typical for tetrameric lac repressor.

Amino Acid Sequence↗
PubMed1976

Isolation of a set of hybrid lac repressors made in vitro between normal lac repressor and its homogeneous tryptic core.

Lactose repressor can be renatured from 8 M guanidine-HCl solution. The renatured repressor is tetrameric and shows DNA binding activity. Thus it becomes possible to obtain hybrid tetramers in vitro between normal repressor and repressor defective in DNA binding by simultaneous denaturation and renaturation. In order to facilitate the separation of the different hybrids, we have used a lac repressor derivative that does not bind DNA, which is missing the amino-terminal 59 residues of the polypeptide chain (homogeneous tryptic core). The hybrids resulting from the mixed renaturation of homogeneous tryptic core and normal repressor can be separated by electrophoresis on Cellogel. The hybrids have been recovered, and a preliminary characterization of their DNA-binding properties is reported.

Bacterial Proteins↗
PubMed1975

Amino-acid sequence of lac repressor from Escherichia coli. Isolation, sequence analysis and sequence assembly of tryptic peptides and cyanogen-bromide fragments.

The lac repressor from Escherichia coli, composed of four identical subunits with a molecular weight of 37160, was carboxymethylated and fragmented by tryptic digestion and cyanogen bromide treatment. Using ion-exchange chromatography, gel filtration and preparative thin-layer electrophoresis and chromatography 29 of the 30 tryptic peptides were isolated in pure form. Direct Edman degradation and the dansyl-Edman technique were used to determine the sequence of the small tryptic peptides. Special emphasis was put on the sequence determination of the six large tryptic fragments which together account for 177 residues, corresponding to 51% of the repressor subunit with its 347 residues. The large tryptic fragments were analyzed after fragmentation with chymotrypsin, thermolysin and dipeptidyl aminopeptidase I. Thus the sequence of all 30 tryptic peptides could be deduced. The complete sequences of all cyanogen bromide fragments were deduced from peptides obtained by tryptic, chymotryptic and thermolytic digestion of the individual fragments and by automated stepwise Edman degradation of lac repressor and of the large cyanogen bromide fragments. The order of the cyanogen bromide fragments was given by overlapping tryptic peptides. The resulting amino acid composition of the monomer is Asp15, Asn11, Thr18, Ser30, Glu14, Gln27, Pro13, Gly22, Ala44, Cys3, Val33, Met9, Ile17, Leu40, Tyr8, Phe4, Trp2, Lys11, His7, Arg19. The sequence of lac repressor shows no similarities with that of other proteins known to bind to DNA or RNA. The N-terminal 55 residues contain two homologous regions. This part of the sequence which is involved in lac operator binding might have been formed by gene duplication.

Amino Acid Sequence↗
PubMed1973

The amino-acid sequence of lac repressor.

The amino-acid sequence of lac repressor from Escherichia coli has been determined. The sequence contains 347 residues in the subunit single peptide chain. It shows no similarities with the sequences of histones or the known part of beta-galactosidase.

Amino Acid Sequence↗
PubMedDate not supplied

Peptides from the conserved ends of the rod domain of desmin disassemble intermediate filaments and reveal unexpected structural features: a circular dichroism, Fourier transform infrared, and electron microscopic study.

Synthetic peptides representing the conserved ends of the rod domain of desmin are shown to disassemble preformed desmin filaments when added in moderate molar excess. This argues for a similar importance of both ends of the rod for filament stability. Recent structural models of intermediate filaments suggest close proximity of the ends and perhaps even an interaction (N. Geisler, J. Schünemann, and K. Weber, 1992, Eur. J. Biochem. 206, 841-852; P. M. Steinert, L. N. Marekov, R. D. B. Fraser, and D. A. D. Parry, 1993, J. Mol. Biol. 230, 436-452). Since the disassembling activity of the peptides, in addition to their sequences, should be related in some way to their secondary structure, we have investigated the structures of a number of related peptides which all arise from the ends of the rod using electron microscopic and spectroscopic methods. All peptides showed the expected alpha-helical structure at low concentrations in the presence of trifluoroethanol, as revealed by circular dichroism. At higher concentrations the peptides showed extensive self-aggregation into various types of filaments. The filaments contain the peptides in beta-sheet conformation as shown by Fourier transform infrared spectroscopy.

Amino Acid Sequence↗