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P Dessen

Publications and source records attributed to P Dessen.

At least 55 records · Page 3Linked to original sources
PubMed1986

Escherichia coli tyrosyl- and methionyl-tRNA synthetases display sequence similarity at the binding site for the 3'-end of tRNA.

Covalent modification of Escherichia coli tyrosyl-tRNA synthetase (TyrRS) by the 2',3'-dialdehyde derivative of tRNATyr (tRNAox) resulted in a time-dependent inactivation of both ATP-PPi exchange and tRNA aminoacylation activities of the enzyme. In parallel with the inactivation, covalent incorporation of approximately 1 mol of [14C]tRNATyrox/mol of the dimeric synthetase occurred. Intact tRNATyr protected the enzyme against inactivation by the tRNA dialdehyde. Treatment of the TyrRS-[14C]tRNATyr covalent complex with alpha-chymotrypsin produced two labeled peptides (A and B) that were isolated and identified by sequence analysis. Peptides A and B are adjacent and together span residues 227-244 in the primary structure of the enzyme. The three lysine residues in this sequence (lysines-229, -234, and -237) are labeled in a mutually exclusive fashion, with lysine-234 being the most reactive. By analogy with the known three-dimensional structure of the homologous tyrosyl-tRNA synthetase from Bacillus stearothermophilus, these lysines should be part of the C-terminal domain which is presumed to bind the cognate tRNA. Interestingly, the labeled TyrRS structure showed significant similarities to the structure around the lysine residue of E. coli methionyl-tRNA synthetase which is the most reactive toward tRNAMetf(ox) (lysine-335) [Hountondji, C., Blanquet, S., & Lederer, F. (1985) Biochemistry 24, 1175-1180].

Amino Acid Sequence↗
PubMed1986

Principle of codification for quick comparisons with the entire biomolecule databanks and associated programs in FORTRAN 77.

We propose a new method for homology search of nucleic acids or proteins in databanks. All the possible subsequences of a specific length in a sequence are converted into a code and stored in an indexed file (hash-coding). This preliminary work of codifying an entire bank is rather long but it enables an immediate access to all the sequence fragments of a given type. With our method a strict homology pattern of twenty nucleotides can be found for example in the Los Alamos bank (GENBANK) in less than 2 seconds. We can also use this data storage to considerably speed up the non-strict homology search programs and to write a program to help in the selection of nucleic acid hybridization probes.

Amino Acid Sequence↗
PubMed1986

Sequence similarities among the family of aminoacyl-tRNA synthetases.

Recent affinity labeling studies have led to the identification of lysine residues at the CCA binding site of tRNA in Escherichia coli methionyl- and tyrosyl-tRNA synthetases. The comparison of the labeled peptides to the known primary structures of the aminoacyl-tRNA synthetases reveals new sequence similarities among this family of enzymes. These similarities include a 'constant' lysine residue whose functional significance is discussed. Moreover, a systematic computer analysis was conducted to search for similarities between the aminoacyl-tRNA synthetases taken as pairs.

Amino Acyl-tRNA Synthetases↗
PubMed1985

Methionyl-tRNA synthetase from E. coli: direct evidence for exchange of protomers in the dimeric enzyme by using deuteration and small-angle neutron scattering.

Direct demonstration of the reversible dissociation of native dimeric methionyl-tRNA synthetase from E. coli has been obtained using small angle neutron scattering and deuterated enzyme. Structural parameters of the fully deuterated dimer are very similar to the hydrogenated one. Analysis of the variations of the intensity and of the radius of gyration of a stoichiometric mixture of the two types of dimer (hydrogenated and deuterated), as a function of D2O content in the solvent, enabled us to characterize an hybrid dimer, having both hydrogenated and deuterated protomers. By separating the contribution of each protomer to the scattering, the radius of gyration of the protomer in situ and the distance between the centers of mass of each protomer in the dimer are determined.

Amino Acyl-tRNA Synthetases↗
PubMed1984

Drug-binding properties of rat alpha 1-foetoprotein. Binding of warfarin, phenylbutazone, azapropazone, diazepam, digitoxin and cholic acid.

As part of an investigation into whether alpha 1-foetoprotein (alpha 1-FP) plays the same transport role in foetal serum as albumin does in the adult, the binding properties of both proteins were compared with respect to the binding of a series of compounds known to be bound by albumin's specific drug-binding sites. The binding of warfarin, phenylbutazone, azapropazone, diazepam, digitoxin and cholic acid by rat alpha 1-FP and serum albumin was studied by equilibrium dialysis at 4 degrees C. Rat alpha 1-FP was shown to have neither albumin's high-affinity site II (diazepam as marker) nor its site III (digitoxin and cholic acid as markers). High-affinity binding by alpha 1-FP was found for the specific markers (warfarin, phenylbutazone, azapropazone) of albumin's drug-binding site I. However, instead of albumin's one high-affinity site/molecule, a mean value of 0.5 site/molecule was obtained with rat alpha 1-FP. Charcoal treatment at neutral pH of rat serum albumin did not affect its measured binding properties, but treatment of the alpha 1-FP led to an increased affinity for warfarin, phenylbutazone and azapropazone without a change in the measured number of sites, indicating competition for binding at this site by (an) endogenous ligand(s). These results are discussed in terms of the structures of the two proteins and with respect to the physiological implications of the differences found.

Animals↗
PubMed1984

Sequence of the initiation factor IF2 gene: unusual protein features and homologies with elongation factors.

The gene for protein synthesis initiation factor IF2 in Escherichia coli, infB, is located downstream from nusA on the same operon. We sequenced about 3 kilobases of DNA beginning within nusA and including the entire infB structural gene plus another 392 bases downstream. This region contains no obvious strong promoter signals, but a possible transcriptional termination or pausing site occurs downstream from infB. The putative initiator codon for IF2 alpha (97,300 daltons) is AUG; that for IF2 beta (79,700 daltons) is GUG, located 471 bases downstream in the same reading frame. The codon usage for IF2 is typical of other highly expressed proteins in E. coli and suggests that IF2 mRNA is efficiently translated. IF2 alpha contains two adjacent regions (residues 104-155 and 167-214) that are rich in alanine and charged amino acids and that show striking periodicities in their sequences. These regions may alternate between flexible and helical conformations, thereby drawing together the NH2-terminal and COOH-terminal globular domains of the factor as IF2 interacts with ribosomes or tRNA. Certain regions of the DNA and protein sequences of IF2 share strong homologies with elongation factor EF-Tu and lesser homology with EF-G. In particular, a region of EF-Tu implicated in GTP binding contains sequences and secondary structure that are conserved in IF2. The homologies indicate that the genes for IF2 and the elongation factors are derived at least in part from a common ancestor.

Amino Acid Sequence↗
PubMed1983

Neutron scattering study of the binding of tRNAPhe to Escherichia coli phenylalanyl-tRNA synthetase.

Escherichia coli phenylalanyl-tRNA synthetase has been characterized by small-angle neutron scattering. In solution (20 mM imidazole hydrochloride, pH 7.6, 10 mM 2-mercaptoethanol, and 0.1 mM ethylenediaminetetraacetic acid), this enzyme has a molecular weight of 227K +/- 20K with a radius of gyration of 48.3 +/- 0.6 A, independent of the presence of MgCl2 up to 50 mM. The change of the scattering upon adding tRNAPhe to the enzyme has been followed with 10 mM MgCl2 present in the buffer. One enzyme molecule is capable of binding two tRNAPhe molecules with affinity constants larger than 10(6) M-1. Parallel titration experiments in 73% 2H2O, close to the matching point of tRNA, show that the RG of the enzyme is not changed by the binding of one or two tRNAPhe molecules. These results are compared with quasi-electric light scattering studies [Holler, E., Wang, C. C., & Ford, N.C., Jr. (1981) Biochemistry 20, 861-867] where the addition of either MgCl2 or tRNAPhe was shown to cause dramatic changes of the apparent translational diffusion constant of phenylalanyl-tRNA synthetase.

Amino Acyl-tRNA Synthetases↗
PubMed1983

Interaction between rat alpha 1 fetoprotein and fluorescent derivatives of estrone in relation to the position and type of the fluorescent label.

A series of derivatives of estrone with fluorescent dyes (dansyl or coumarin) coupled at different positions on the steroid molecule, have been synthesized. These derivatives were tested for their quantum yields and their binding properties were determined with respect to rat alpha 1 fetoprotein. Derivatives at C-3 (estrone-3-hemisuccinate-dansyl-cadaverine and estrone-3-dansyl) compete with estrone for binding to the fetal protein; however derivates of estrone at C-6 (estrone-6-carboxymethyloxime-dansyl-cadaverine) and at C-17 (estrone-17-carboxymethyloxime-coumarine, estrone-17-carboxymethyloxime-dansyl-cadaverine and estrone-17-dansyl-hydrazine) compete poorly or not at all. The association constant of the radioactive derivative estrone-3-dansyl [3H] with rat alpha 1 fetoprotein was measured directly: the same number of high affinity binding sites (0.6) as that for estrone was found with an apparent association constant of 3.7 X 10(6) M-1. In addition to the high affinity binding sites, a low affinity class of binding sites was found which corresponds to the binding of the dansyl fraction of the fluorescent steroid derivative.

Animals↗
PubMed1982

Sequence of a 1.26-kb DNA fragment containing the structural gene for E.coli initiation factor IF3: presence of an AUU initiator codon.

The nucleotide sequence of a 1.26-kb pair DNA fragment containing the structural gene for Escherichia coli initiation factor IF3 has been determined. An open reading frame of 540 nucleotides is found at the position predicted by genetic studies. The amino-acid sequence deduced from the DNA sequence accounts for a molecular weight 20 530. The important feature of the coding DNA sequence is the presence of AUU as the translational initiator codon. It is 11 bases downstream of the center of a GGAGG sequence, which can strongly pair with the sequence CCUCC near the 3' terminus of 16S rRNA. The primary DNA sequence in the region of the AUU initiator codon and its role in compensating a reduced codon-anticodon interaction in initiation complex formation are discussed.

Amino Acid Sequence↗
PubMed1980

Proteolytic cleavage of methionyl transfer ribonucleic acid synthetase from Bacillus stearothermophilus: effects on activity and structure.

Methionyl-tRNA synthetase from Bacillus stearothermophilus, a dimer of molecular weight 2 X 85K, is converted by limited subtilisin digestion into a fully active monomeric fragment of molecular weight 64K. The reversible methionine activation reaction of these enzymes was followed through the variation of the intensity of their trypotophan fluorescence. Equilibrium and stopped-flow experiments show that the rate and mechanism for adenylate formation supported by the monomeric derivative are undistinguishable from those of each adenylating site of the native dimeric enzyme. In contrast, the rate of tRNA aminoacylation is improved upon limited proteolysis of the native enzyme. This behavior can be related to the anticooperativity of the binding of tRNA molecules to native dimeric enzyme. Accordingly, at 25 degrees C, the dimer might behave as a half-of-the-sites enzyme with only one active tRNA site at a time, compared to two after limited proteolysis with consequent irreversible disociation into two 64K fragments. Another modified form of the enzyme is obtained through limited tryptic digestion. This derivative is completely devoid of activity although its molecular weight under nondenaturating conditions remains undistinguishable from that of the 64K fragment generated by subtilisin. Denaturation reveals that this tryptic derivative is composed of two subfragments with molecular weights of 33K and 29K, respectively. The same fragments may also be directly obtained through limited tryptic digestion of the subtilsic fragment. Interestingly, although trypsin treatment has abolished the activity of the enzyme, fluorescence studies demonstrate that the ATP and methionine binding sites have remained intact. It is shown that the effect of the internal cut made by trypsin into the active 64K fragment has been to considerably depress the "coupling" between the methionine and nucleotide binding sites. Finally, the rate of inactivation of the enzyme by trypsin is observed to be substantially decreased by in situ synthetized methionyl adenylate but not by tRNA. These properties and others are discussed in relation to the problem of its significance of repeating sequences and structural "domains" within the class of aminoacyl-tRNA synthetases.

Adenosine Triphosphate↗