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A Tramontano

Publications and source records attributed to A Tramontano.

At least 91 records · Page 5Linked to original sources
PubMed1991

Members of the zinc finger protein gene family sharing a conserved N-terminal module.

We report the isolation of human members of a sub-family of structurally related finger protein genes. These potentially encode polypeptides containing finger motifs of the Krüppel type at the C-terminus, and a conserved amino acid module at the N-terminus; because of its invariant location the latter is referred to as finger preceding box (FPB). The FPB, detected also in previously described finger proteins from human, mouse and Xenopus, extends over approximately 65 amino acids and appears to be composed of two contiguous modules: FPB-A (residues 1-42) and FPB-B (residues 43-65). The latter is absent in some of the members analyzed. Elements A and B and the zinc finger domain are encoded by separate exons in the ZNF2 gene, a human member of this sub-family. The positioning of introns within this gene is remarkable. One intron flanks and a second interrupts the first codon of the FPB-A and FPB-B modules, respectively. A third intron occurs a few nucleotides downstream of FPB-B marking its separation from the remainder of the coding sequences. This organization, together with the absence of FPB-B in some cDNAs, supports the hypothesis that mRNAs encoding polypeptides that include one, both or none of the FPB-A and FPB-B modules may be assembled through alternative splicing pathways. Northern analyses showed that members of this sub-family are expressed as multiple transcripts in several cell lines. The sequences of distinct cDNAs homologous to the ZNF2 gene indicate that alternative splicing events adjoin either coding or non coding exons to the FPB sequences.

Amino Acid Sequence↗
PubMed1990

Chemical equilibrium at an antibody binding site: catalytic efficiency defined by a Haldane relationship.

This paper describes the first study of a reaction catalyzed in both forward and reverse directions by an antibody. The rates of reversible addition of sulfite to 6-hydroxy-3H-xanthen-3-one (1) and their pH dependence are influenced by a monoclonal antibody to a fluorescein hapten. The antibody is presumed to recognize the two substrates in a chemically productive prereaction complex with a geometry similar to that between the carboxyl group and the xanthenyl group in fluorescein. Equilibria are determined for the reactions in solution and at the antibody combining site from rate constants for the forward and reverse reactions. The solution equilibrium lies in favor of adduct with KA = x 10(7) M-1 at pH 7.5. The corresponding equilibrium at the combining site is much smaller (KAb = 8), and this is attributed to the preferential binding of substrates 1 and sulfite. The magnitude of differential stabilization is determined from the equilibrium constants (2 x 10(6) M-1). Observed rates at low molar standard state are compared to reveal the acceleration of the process at the antibody combining site versus the bimolecular solution reaction. In the associative direction, the rate factor of 1200 may largely be attributed to the entropy savings for the reaction on the catalyst. This analysis serves to illustrate the potential for catalysis by antibodies due from recognition of structural differences between substrates of different energy on a reaction coordinate and suggests a strategy for inducing antibody catalysts that does not presume knowledge of catalytic mechanisms or transition-state structure.

Antibodies, Monoclonal↗
PubMed1990

Framework residue 71 is a major determinant of the position and conformation of the second hypervariable region in the VH domains of immunoglobulins.

Analysis of the immunoglobulins of known structure reveals systematic differences in the position and main-chain conformation of the second hypervariable region of the VH domain (H2). We show that the major determinant of the position of H2 is the size of the residue at site 71, a site that is in the conserved framework of the VH domain. It is likely that for about two thirds of the known VH sequences the size of the residue at this site is also a major determinant of the conformation of H2. This effect can override the predisposition of the sequence, as in the case of the H2 loop of J539, which is an exception to the rules relating sequence and conformation of short hairpin loops. Understanding the relationship between the residue at position 71 and the position and conformation of H2 has applications to the prediction and engineering of antigen-binding sites of immunoglobulins.

Amino Acid Sequence↗
PubMed1990

VARIANT: a store and retrieval system for human haemoglobin variants.

VARIANT, a FORTRAN 77 computer system for storing and retrieving the amino acid replacements in human haemoglobin variants on a multicriterion basis, is presented. The system is transportable and user-friendly, can easily be expanded and can also be used to create a database of the amino acid replacements in families of homologous proteins, becoming an efficient tool for evolutionary studies.

Amino Acids↗
PubMed1990

Structural definition by antibody engineering of an idiotypic determinant.

Using computer-aided techniques for predicting molecular structure, we constructed an atomic model of the variable domain of a murine anti-thyroglobulin antibody whose immunodominant idiotypic determinant (Id62) was mapped by site-directed mutagenesis and immunochemical analysis. We previously showed that under experimental conditions this idiotype activates anti-idiotypic B cells and T cells, and modulates the response to thyroglobulin in mice. Because idiotype interactions are considered of physiological importance for immune regulation, we studied this idiotype as a model to understand the relationship between function and structure. To determine the contribution of heavy- and light-chain variable domains to the idiotype structure, we constructed chimeric expression vectors and introduced them into the (non-secreting) P3X63Ag8.653 myeloma cell line. Mutants of the heavy-chain variable domain were obtained by site-directed mutagenesis and transfected into the murine (lambda 1) light-chain producer J558L cell line. The expressed proteins were purified from culture supernatants of transfected cells and characterized. We provide evidence that the third hypervariable loop (D region) of the heavy-chain variable domain is the structural correlate of the idiotypic determinant of this autoantibody and is independent from the nature of the associated light chain. Substitution of residues of the first and second complementarity-determining regions do not affect idiotype expression. The results described here are discussed in relation to our understanding, at a molecular level, of the interaction of idiotopes with B- and T-cell compartments.

Amino Acid Sequence↗
PubMed1989

Structural determinants of the conformations of medium-sized loops in proteins.

Loops are integral components of protein structures, providing links between elements of secondary structure, and in many cases contributing to catalytic and binding sites. The conformations of short loops are now understood to depend primarily on their amino acid sequences. In contrast, the structural determinants of longer loops involve hydrogen-bonding and packing interactions within the loop and with other parts of the protein. By searching solved protein structures for regions similar in main chain conformation to the antigen-binding loops in immunoglobulins, we identified medium-sized loops of similar structure in unrelated proteins, and compared the determinants of their conformations. For loops that form compact substructures the major determinant of the conformation is the formation of hydrogen bonds to inward-pointing main chain atoms. For loops that have more extended conformations, the major determinant of their structure is the packing of a particular residue or residues against the rest of the protein. The following picture emerges: Medium-sized loops of similar conformation are stabilized by similar interactions. The groups that interact with the loop have very similar spatial dispositions with respect to the loop. However, the residues that provide these interactions may arise from dissimilar parts of the protein: The conformation of the loop requires certain interactions that the protein may provide in a variety of ways.

Binding Sites↗
PubMed1988

A transportable interactive package for the statistical analysis and handling of sequence data.

A real-time, machine-independent software for the analysis and manipulation of sequence data is described. The implemented system allows users to read in sequence data from existing data bases, and to edit, manipulate, analyse and align them. It is easy to use and is suitable for routine investigations and to modify sequences in order to create a user customized data base.

Algorithms↗
PubMed1987

A stereospecific cyclization catalyzed by an antibody.

A monoclonal antibody elicited by a transition-state analog that is representative of an intramolecular six-membered ring cyclization reaction acted as a stereospecific, enzyme-like catalyst for the appropriate substrate. Formation of a single enantiomer of a delta-lactone from the corresponding racemic delta-hydroxyester was accelerated by the antibody by about a factor of 170, which permitted isolation of the lactone in an enantiomeric excess of about 94 percent. This finding demonstrates the feasibility of catalytic-antibody generation for chemical transformations that require stereochemical control.

Antibodies, Monoclonal↗
PubMed1986

Catalytic antibodies.

Monoclonal antibodies elicited to haptens that are analogs of the transition state for hydrolysis of carboxylic esters behaved as enzymic catalysts with the appropriate substrates. These substrates are distinguished by the structural congruence of both hydrolysis products with haptenic fragments. The haptens were potent inhibitors of this esterolytic activity, in agreement with their classification as transition state analogs. Mechanisms are proposed to account for the different chemical behavior of these antibodies with two types of ester substrates. The generation of an artificial enzyme through transition state stabilization by antibodies was thus demonstrated. These studies indicate a potentially general approach to catalyst design.

Antibodies, Monoclonal↗
PubMed1986

Probability of coding of a DNA sequence: an algorithm to predict translated reading frames from their thermodynamic characteristics.

An algorithm to determine the probability that a reading frame codifies for a protein is presented. It is based on the results of our previous studies on the thermodynamic characteristics of a translated reading frame. We also develop a prediction procedure to distinguish between coding and non-coding reading frames. The procedure is based on the characteristics of the putative product of the DNA sequence and not on periodicity characteristics of the sequence, so the prediction is not biased by the presence of overlapping translated reading frames or by the presence of translated reading frames on the complementary DNA strand.

Amino Acid Sequence↗
PubMed1986

Chemical reactivity at an antibody binding site elicited by mechanistic design of a synthetic antigen.

Monoaryl phosphonate esters, designated as analogs of the transition state in the hydrolysis of carboxylic esters, were synthesized and used as haptens to generate specific monoclonal antibodies. Some of these antibodies react with cognate aryl carboxylic esters to release a fluorescent alcohol. The reaction appears to be stoichiometric; however, the activity is slowly regenerated under alkaline conditions or by treatment with hydroxylamine. Specificity is rigorous for esters of p-trifluoroacetamidophenylacetic acid, demonstrating a structural correspondence with the phosphonate hapten. Saturation kinetics are observed and kinetic parameters (kmax, Vmax, and Km) are reported. The haptenic phosphonate is a competitive inhibitor of the reaction (Ki, 35 nM); whereas the carboxylate product of ester hydrolysis is a less effective inhibitor (Ki, ca. 7500 nM). Chemical modification of side chain groups in the protein show a partial reduction in activity on acylation of lysine or nitration of tyrosine and a dramatic quenching upon modification of histidine. The evidence is discussed in terms of a mechanism in which amino acids of the antibody combining site participate in nucleophilic and/or general base catalysis. The properties of this system suggest that it is an example of enzymic transacylation where a deacylation step is not catalyzed. The possibility of deriving enzymic function from immunological specificity through this approach is advanced.

Acylation↗
PubMed1985

Determination of the autocorrelation orders of proteins.

A statistical analysis of the autocorrelation characteristics of active polypeptides has been carried out by means of the correlogram method. It is shown that the amino acid sequences of the analysed proteins, considered as a whole, are autocorrelated and that the correlograms characterize each protein reflecting its three-dimensional structure.

Amino Acid Sequence↗
PubMed1984

Statistical evaluation of the coding capacity of complementary DNA strands.

Two independent methods are used to evaluate the protein-coding information content in different classes of DNA sequences. The first method allows to evaluate the statistical relevance of finding unidentified reading frames, longer than 100 codons, on both DNA strands of: a) 117 DNA sequences that code for 142 nuclear proteins; b) 39 stable RNA coding sequences and c) 36 other DNA sequences which include regulatory and as yet unknown function sequences. The finding of 50 reading frames longer than 100 codons (complementary inverted proteins or c.i.p. genes) located on the DNA strand complementary to the protein-coding one is drastically in excess of the number predicted by chance alone. An independent method (testcode) applied to c.i.p. gene sequences, which assigns the probability of coding to a given sequence, predicts that more than 50% of these genes are translated in a functional product. These analyses indicate the existence of a new class of protein-coding genes, located on the DNA sequences complementary to the protein-coding DNA strand.

Animals↗
PubMed1982

Thermodynamic approach to a possible theory of the evolution of a genetic code.

One of the distinctive features of a biological system is its remoteness from equilibrium: for the treatment of the information exchanged by these systems with the external world the concept of information amount is not sufficient because for nonequilibrium systems we need further specifications about information, i.e. its value. We defined the information value for a genetic code. The comparison of the results we had applying such a definition to the three genetic codes found in nature shows that our definition described the system of transmission and reception of information in a biological organism very well and that the information value of a genetic code can be considered as an index of its efficiency, i.e. of its ability to minimize the effects of a mutation of the genotype on the phenotype. Otherwise, our results show that the information value, and hence the efficiency of the three known codes is the same and suggest that the prerequisite of the evolution of the codes is the preservation of this value.

Biological Evolution↗