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Biomedical subjects

R A Lerner

Publications and source records attributed to R A Lerner.

At least 55 records · Page 3Linked to original sources
PubMed1995

Reactive immunization.

For almost 200 years inert antigens have been used for initiating the process of immunization. A procedure is now described in which the antigen used is so highly reactive that a chemical reaction occurs in the antibody combining site during immunization. An organophosphorus diester hapten was used to illustrate this concept coined "reactive immunization." The organophosphonate recruited chemical potential from the immune response that resembled the way these compounds recruit the catalytic power of the serine hydrolases. During this recruitment, a large proportion of the isolated antibodies catalyzed the formation and cleavage of phosphonylated intermediates and subsequent ester hydrolysis. Reactive immunization can augment traditional immunization and enhance the scope of catalytic antibody chemistry. Among the compounds anticipated to be effective are those that contain appropriate reactive functionalities or those that are latently reactive, as in the mechanism-based inhibitors of enzymes.

Animals↗
PubMed1995

Efficient aldolase catalytic antibodies that use the enamine mechanism of natural enzymes.

Antibodies that catalyze the aldol reaction, a basic carbon-carbon bond-forming reaction, have been generated. The mechanism for antibody catalysis of this reaction mimics that used by natural class I aldolase enzymes. Immunization with a reactive compound covalently trapped a Lys residue in the binding pocket of the antibody by formation of a stable vinylogous amide. The reaction mechanism for the formation of the covalent antibody-hapten complex was recruited to catalyze the aldol reaction. The antibodies use the epsilon-amino group of Lys to form an enamine with ketone substrates and use this enamine as a nascent carbon nucleophile to attack the second substrate, an aldehyde, to form a new carbon-carbon bond. The antibodies control the diastereofacial selectivity of the reaction in both Cram-Felkin and anti-Cram-Felkin directions.

Acetone↗
PubMed1995

From molecular diversity to catalysis: lessons from the immune system.

By combining the enormous molecular diversity of the immune system with basic mechanistic principles of chemistry, one can produce catalytic antibodies that allow control of reactions in ways heretofore not possible. Mechanistic and structural studies of these antibodies are also providing insights into important aspects of enzymatic catalysis and the evolution of catalytic function. Moreover, the ability to rationally direct the immune response to generate selective catalysts for reactions ranging from pericyclic and redox reactions to cationic rearrangement reactions underscores the chemical potential of this and other large combinatorial libraries.

Amino Acid Sequence↗
PubMed1995

Chemical characterization of a family of brain lipids that induce sleep.

A molecule isolated from the cerebrospinal fluid of sleep-deprived cats has been chemically characterized and identified as cis-9,10-octadecenoamide. Other fatty acid primary amides in addition to cis-9,10-octadecenoamide were identified as natural constituents of the cerebrospinal fluid of cat, rat, and human, indicating that these compounds compose a distinct family of brain lipids. Synthetic cis-9,10-octadecenoamide induced physiological sleep when injected into rats. Together, these results suggest that fatty acid primary amides may represent a previously unrecognized class of biological signaling molecules.

Animals↗
PubMed1995

Encoded reaction cassette for the highly sensitive detection of the making and breaking of chemical bonds.

A reaction cassette has been designed for the highly sensitive detection of the making and breaking of chemical bonds. The system is envisioned as a companion device to be used in the search for antibody and other novel catalysts. The cassette also may have important clinical applications in the design of diagnostic reagents. In its fully encoded format, this methodology is capable of both detecting and decoding chemical events.

Antibodies↗
PubMed1995

Catalytic antibodies: evolution of protein function in real time.

Natural selection of enzyme function has evolved over millions to billions of years, whereas antibody induction operates over a period of weeks. If one considers how new protein functions are generated, one sees that both the immune system and natural selection have powerful methods for the generation of diversity. In this treatise we discuss antibody catalysis from an evolutionary standpoint as we consider how the elicitation of a catalytic antibody can be a useful tool for exploring the nature of biological catalysis.

Amides↗
PubMed1995

Random mutagenesis of staphylococcal nuclease and phage display selection.

Multiple cycles of mutagenesis and phage display selection have been investigated as a method for obtaining enzymes with altered catalytic properties. A library of staphylococcal nuclease mutants displayed on phage was created by error-prone PCR mutagenesis and selected for binding to thymidine- or guanosine-containing substrate analogs. After discarding non-binders, the binding mutants were then subjected to further mutagenesis and selection rounds. After four mutagenesis and selection cycles, the catalytic properties of some of the resulting nucleases were studied and one nuclease with nine accumulated mutations was found to have a two-fold reduction in kcat for DNA hydrolysis, but a two-fold increase in kcat/Km for hydrolysis of a thymidine containing small molecule substrate. The possibility of this technique for in vitro evolution of enzyme properties is discussed.

Amino Acid Sequence↗
PubMed1994

Cerebrodiene: a brain lipid isolated from sleep-deprived cats.

We report the isolation of a heretofore unrecognized brain lipid that is present in cerebrospinal fluid of sleep-deprived cats. The molecule appears to be a long-chain base structurally related to sphingosine and sphinganine in which a second unsaturated bond has been introduced. An increase in the degree of unsaturation of a key membrane component is expected to have important physiological consequences.

Animals↗
PubMed1994

Antibody catalyzed cationic cyclization.

Two major goals for the design of new catalysts are the facilitation of chemical transformations and control of product outcome. An antibody has been induced that efficiently catalyzes a cationic cyclization in which an acyclic olefinic sulfonate ester substrate is converted almost exclusively (98 percent) to a cyclic alcohol. The key to the catalysis of the reaction and the restriction of the product complexity is the use of antibody binding energy to rigidly enforce a concerted mechanism in accord with the design of the hapten. Thus, the ability to direct binding energy allows the experimenter to dictate a reaction mechanism which is an otherwise difficult task in chemistry. New catalysts for cationic cyclization may be of general use in the formation of carbon-carbon and carbon-heteroatom bonds leading to multi-ring molecules including steroids and heterocyclic compounds.

Animals↗
PubMed1994

Direct selection for a catalytic mechanism from combinatorial antibody libraries.

Semisynthetic combinatorial antibody library methodology in the phage-display format was used to select for a cysteine residue in complementarity-determining regions. Libraries were panned with an alpha-phenethyl pyridyl disulfide that undergoes disulfide interchange. Out of 10 randomly picked clones, two contained an unpaired cysteine, one of which was studied. The antibody catalyzed the hydrolysis of the corresponding thioester where the electrophilic carbonyl occupies the three-dimensional space that was defined by the reactive sulfur atom during selection. The reaction operates by covalent catalysis. Although the steady-state rate enhancement relative to the activated thiol ester substrate is modest, hydrolysis of the acylated cysteine intermediate is remarkably efficient with a catalytic advantage of about four orders of magnitude. The results suggest that iterative mechanism-based selection procedures can recapitulate the enzymatic mechanisms refined through evolution.

Amino Acid Sequence↗
PubMed1993

Selection of human anti-hapten antibodies from semisynthetic libraries.

Semisynthetic human Fab libraries were constructed, displayed on the surface of filamentous phage and selected for binding to three hapten conjugates. A number of Fabs were isolated and characterized with respect to affinity and specificity. Fabs exhibited affinities of between 80 and 29 nM, as determined by surface plasmon resonance, for the conjugate on which they were selected. Conservation of Asp101 in the third heavy-chain complementarity determining region (HCDR3) appears to be important in the construction of synthetically diverse repertoires.

Amino Acid Sequence↗
PubMed1993

Control of the exo and endo pathways of the Diels-Alder reaction by antibody catalysis.

Catalytic antibodies that control the reaction pathways of the Diels-Alder cycloaddition have been generated. One antibody catalyzes the favored endo and the other the disfavored exo pathway to yield the respective cis and trans adducts in enantiomerically pure form. A comparison of the x-ray structure of the hapten with the calculated geometry of the transition structure showed that [2.2.2] bicyclic compounds are excellent mimics of the transition state of the Diels-Alder reaction. To achieve catalysis and the high degree of stereoselectivity shown here, the antibody must simultaneously control the conformation of the individual reactants and their relation to each other. In the case of the disfavored process, binding energy must be used to reroute the reaction along a higher energy pathway. The rerouting of reaction pathways has become a major focus of antibody catalysis and other disfavored reactions can be expected to be catalyzed so long as the energy barrier is not extreme. The energy requirements needed for absolute control of all of the stereoisomers of many Diels-Alder reactions fall in the energy range (approximately 20 kilocalories per mole) deliverable by antibody binding.

Acrylamide↗
PubMed1993

Direct selection of antibodies that coordinate metals from semisynthetic combinatorial libraries.

An iterative strategy for the selection of catalytic metalloantibodies is described. The first stage of this strategy is validated by the selection of semisynthetic antibodies that coordinate a variety of different metal ions and the metal oxide magnetite. These results have implications not only for the development of catalytic metalloantibodies but also for the development of reagents for magnetic resonance imaging, delivery of radioisotopes, and purification of recombinant proteins.

Amino Acid Sequence↗
PubMed1993

Catalytic antibodies: the concept and the promise.

Catalytic antibodies both label antigenic targets and perform enzymatic functions. Since the conceivable specificities are virtually limitless, the potential of such tools is great. They can, for example, be engineered to destroy specific pathogens or tumor cells, to protect normal cells from chemotoxicity, or to activate prodrugs at the target site.

Antibodies, Catalytic↗
PubMed1993

Antibody catalysis of a disfavored chemical transformation.

Organic reactions are often limited by stereoelectronic constrains that appear along the reaction coordinate. An antibody has been generated that overcomes these constraints and catalyzes a highly disfavored chemical transformation. The antibody facilitates the difficult 6-endo-tet ring closure of an epoxy-alcohol to form a tetrahydropyran. The catalyzed process is in formal violation of what has become known as Baldwin's rules for ring-closure reactions. In addition to controlling the regiochemistry of the disfavored cyclization reaction, these catalytic antibodies resolve enantiomeric substrates to afford a stereochemically pure product. The principles demonstrated in this study may be applicable to other disfavored chemical processes.

Antibodies↗