Can enzymes adopt a self-inhibited form? Results of x-ray crystallographic studies of chymosin.

Chymosin molecules in the crystal lattice have Tyr77 occluding the S1/S3 substrate binding pockets suggesting that the enzyme is self-inhibited. An analysis of this structure in conjunction with its comparison with pepsin has shown that this is most probably an intrinsic property of the enzyme. It also indicates that chymosin's substrate specificity may be dependent upon the ability of the substrate to displace the tyrosine ring from the binding pockets.

Trivaline 'catalyzes' 5'-pdGTT oligomerization in solution.

We have found that the 5'-pdGTT molecules at a concentration of 10(-4) M are oligomerized in solution in the presence of 10(-4) M tripeptide-(L-Val)3-NH-NH-DNS.CF3COOH and the condensation reagents (carbodiimide and imidazole). Oligonucleotides not less than 12 bases long were formed in the yield which was over 15%.

Southern molecular hybridization experiments with parallel complementary DNA probes.

We have detected the specific binding in Southern blot hybridization experiments of both complementary antiparallel and parallel 40 bp synthetic DNA probes, corresponding to a cloned Drosophila DNA fragment. The highly cooperative annealing and melting were observed in solution with these probes, which are complementary in the same direction and possess 17 GC pairs. The binding of ethidium bromide is indicative of formation of a perfect parallel DNA duplex.

Structural organization of kinetoplast DNA and its compaction in the in vitro model system.

Electron microscopic studies of Leishmania gymnodactyli cells lysed at hypotonic conditions showed that the structures identified as kinetoplast DNA have the appearance of loose accumulations of crossed and sometimes branched rod-like structures 100 to 200 nm long and 20 nm thick. The compaction of isolated kinetoplast DNA (kpDNA) caused by interaction with synthetic tripeptide--dansylhydrazide trivaline--was also studied. The analysis of the structures arising at different steps of compaction showed that the minicircles are compacted forming rod-like structures where minicircle double-stranded DNA segments are closely associated side by side in a manner which was earlier described for initial compaction stages of "triple rings".

General features of chromosome substitutions in Triticum aestivum x T. timopheevii hybrids.

Based on a C-banded chromosome analysis of Triticum aestivum x T. timopheevii hybrid lines, we developed a classification of the A(t) and G genome chromosomes that agrees with the standard genetic nomenclature of T. aestivum chromosomes.

Parallel double stranded helices and the tertiary structure of nucleic acids.

Thermal denaturation of four oligonucleotides, viz. 3'-d(AT)5pO(CH2)6Opd(AT)5-3'(par(AT], 3'-d(AT)5pO(CH2)6Opd(AT)5-5'(anti(AT],3'-d(A)10pO(CH2) 6Op(T)10-3'(par(A-T], and 3'-d(A)10pO(CH2)6Opd(T)10-5' (anti(A-T], was studied in 0.01 M phosphate buffer, pH 7, in the presence of 0.

Properties of promoter regions of mdg1 Drosophila retrotransposon indicate that it belongs to a specific class of promoters.

A sequence 30 bp downstream from the start site of the Drosophila melanogaster retrotransposon mdg1 is shown to be responsible for correct and precise initiation of mdg1 RNA synthesis in combination with the RNA start-site sequence TCAGTT. A sequence-specific DNA binding protein is demonstrated to interact with the +30 sequence, and the efficient binding of this factor is necessary for in vivo transcriptional activity of the plasmid constructs containing mdg1 promoter fragments. The nucleotides -8/+34 of mdg1 represent a minimal promoter which is able to provide correct initiation of transcription by RNA polymerase II at basal levels.

Sequence-specific cleavage of double-stranded DNA caused by X-ray ionization of the platinum atom in the Pt-bis-netropsin--DNA complex.

An analog of the antibiotic netropsin containing two netropsin-like fragments linked covalently via a platinum atom has been synthesized. DNase I and hydroxyl radical footprinting studies have shown that this compound binds at selective sites on a DNA restriction fragment with a known nucleotide sequence. After X-ray irradiation of Pt-bis-netropsin--DNA complexes a platinum-mediated cleavage of DNA is observed at specific DNA sites.

Allelic variants of rearranged immunoglobulin heavy and light chain genes in hybridoma PTF-02 and parent myeloma.

The hybridoma PTF-02 secretes an antibody against pig transferrin. Rearranged genes for heavy and light immunoglobulin chains have been studied in the genomes of this hybridoma and in the parent myeloma P3-X63.Ag8.

A method for DNA sequencing by hybridization with oligonucleotide matrix.

A new technique of DNA sequencing by hybridization with oligonucleotide matrix (SHOM) which could also be applied for DNA mapping and fingerprinting, mutant diagnostics, etc., has been tested in model experiments. A dot matrix was prepared which contained 9 overlapping octanucleotides (8-mers) complementary to a common 17-mer.

Four-stranded DNA helices: conformational analysis of regular poly(dT).poly(dA).poly(dA).poly(dT) helices with various types of base binding.

The paper presents results obtained in conformational analysis of homopolymeric four-stranded poly(dT).poly(dA).poly(dA).

Interaction of acetyl-CoA fragments with rat liver acetyl-CoA carboxylase.

The interaction of acetyl-CoA fragments with rat liver acetyl-CoA carboxylase has been studied. Dephosphorylated acetyl-CoA did not actually differ from acetyl-CoA in its substrate properties. Non-nucleotide analogues of the substrate, S-acetylpantatheine and it's 4'-phosphate, also possess substrate properties (Vmax = 1.

Linear dichroism studies of tryptophanase and its quasisubstrate complexes oriented in polyacrylamide gel.

Tryptophanase from Escherichia coli was oriented in a compressed slab of polyacrylamide gel and its linear dichroism (LD) and absorption spectra have been measured. The free enzyme displays four LD bands at 305, 340, 425 and 490 nm. Two bands at 340 and 425 nm belong to the internal coenzyme-lysine aldimine.

Inactivation of bacteriophage T7 DNA-dependent RNA polymerase by 5'-p-fluorosulfonylbenzoyladenosine. Identification of the modification site and the effect of the modification on enzyme action.

Bacteriophage T7 RNA polymerase was covalently modified by 5'-[4-fluorosulfonyl)benzoyl]adenosine (4-FSO2BzAdo). The modified enzyme lacks the ability to catalyze RNA synthesis from the phi 10 promoter of bacteriophage T7; both promoter and GTP binding being markedly decreased. The mild hydrolysis of the ester bond of 4-FSO2BzAdo within the covalent enzyme-inhibitor complex restores the RNA synthesis at a lower rate.

X-ray analysis of 2',3'-lyxoanhydrothymidine, a conformationally restricted inhibitor of retroviral reverse transcriptases.

2',3'-Lyxoanhydrothymidine (LAT), a conformationally restricted inhibitor of retroviral reverse transcriptases, has been studied by X-ray analysis. The unit cell contains two crystallographically independent molecules A and B. Their sugar moieties have an identical structure: an 04'-endo pucker of the furanose cycle and a trans conformation about the exocyclic C4'-C5' bond.

Unexpected close relationship between the large nonvirion proteins of filamentous potexviruses and spherical tymoviruses.

The amino acid sequences of the large proteins of potexviruses and tymoviruses were aligned. It was shown that three domains of these proteins display significant similarity between the two virus groups. In contrast to central (putative NTPase-helicase) and C-terminal (putative polymerase) domains, the conservative N-terminal domain of the potexvirus/tymovirus large protein displayed no obvious similarity to the respective regions of the large proteins of other Sindbis-like viruses.

Affinity labelling of rat liver acetyl-CoA carboxylase by a 2',3'-dialdehyde derivative of ATP.

The interaction of rat liver acetyl-CoA carboxylase with a 2',3'-dialdehyde derivative of ATP (oATP) has been studied. The degree of the enzyme inactivation has been found to depend on the oATP concentration and the incubation time. ATP was proved to be the only substrate which protected the inactivation.

T7 RNA polymerase: use of limited proteolysis for the study of enzyme's interaction with substrates and inhibitors.

The specific limited trypsinolysis of bacteriophage T7 RNA polymerase (T7RP) was performed in the presence of various components of the polymerase reaction and some GTP-analogs--irreversible inhibitors of the enzyme. The differences in the rate and sites of proteolysis were observed. Basing on the data obtained the role of the N-terminal domain of the T7RP in the interaction with promoter-containing template is proposed.

An oligonucleotide hybridization approach to DNA sequencing.

We have proposed a DNA sequencing method based on hybridization of a DNA fragment to be sequenced with the complete set of fixed-length oligonucleotides (e.g., 4(8) = 65,536 possible 8-mers) immobilized individually as dots of a 2-D matrix [(1989) Dokl.

Pyrophosphate analogues in pyrophosphorolysis reaction catalyzed by DNA polymerases.

It is demonstrated here that rat liver DNA polymerase beta catalyzes the pyrophosphorolysis reaction with pyrophosphate (PPi) and its analogues. The substrate specificity of the PPi-binding site of several DNA polymerases was investigated. It was discovered that the ability of DNA polymerases to utilize PPi analogues instead of PPi in the pyrophosphorolysis reaction was markedly restricted.