Biomedical subjects

K P Mintz

Publications and source records attributed to K P Mintz.

At least 19 recordsLinked to original sources

PubMed2000

impA, a gene coding for an inner membrane protein, influences colonial morphology of Actinobacillus actinomycetemcomitans.

Directed mutagenesis of a gene coding for a membrane protein of the periodontopathogen Actinobacillus actinomycetemcomitans was achieved by conjugation. The gene was disrupted by insertion of an antibiotic cassette into a unique endonuclease restriction sequence engineered by inverse PCR. The disrupted gene was cloned into a conjugative plasmid and transferred from Escherichia coli to A. actinomycetemcomitans. The allelic replacement mutation resulted in the loss of a 22-kDa inner membrane protein. The loss of this protein (ImpA) resulted in changes in the outer membrane protein composition of the bacterium. Concurrent with the mutation in impA was a change in the pattern of growth of the mutant bacteria in broth cultures. The progenitor bacteria grew as a homogeneous suspension of cells compared to a granular, autoaggregating adherent cell population described for the mutant bacteria. These data suggest that ImpA may play a regulatory role or be directly involved in protein(s) that are exported and associated with colony variations in A. actinomycetemcomitans.

Aggregatibacter actinomycetemcomitans↗

PubMed1999

Binding of the periodontal pathogen Actinobacillus actinomycetemcomitans to extracellular matrix proteins.

The interaction of Actinobacillus actinomycetemcomitans, an important pathogen implicated in juvenile and adult periodontitis, with collagenous and noncollagenous proteins of the extracellular matrix was investigated. A. actinomycetemcomitans SUNY 465 bound to immobilized type I, II, III and V but not type IV collagen. Binding to immobilized collagen was saturable and concentration dependent. This interaction could not be inhibited by soluble collagen, suggesting that binding was dependent on a specific collagen conformation. Bacteria grown anaerobically exhibited decreased collagen-binding activity as compared with organisms grown acrobically. Bacterial outer membrane proteins were essential for binding to collagen. A actinomycetemcomitans SUNY 465 also bound to immobilized fibronectin. In contrast, bacteria did not bind to fibrinogen, bone sialoprotein, alpha 2-HS glycoprotein or albumin. The mechanism of the interaction with fibronectin was more complex, possibly involving both protein and nonproteinaceous components. The majority of other A. actinomycetemcomitans strains tested bound to extracellular matrix proteins in a manner similar to SUNY 465 but with minor variation. These results demonstrate that A. actinomycetemcomitans binds to proteins found in connective tissue. The interaction with extracellular matrix proteins may contribute to the virulence of this pathogen at oral and extraoral sites of infection.

Aggregatibacter actinomycetemcomitans↗

PubMed1999

Virulence factors of Actinobacillus actinomycetemcomitans.

A. actinomycetemcomitans has clearly adapted well to its environs; its armamentarium of virulence factors (Table 2) ensures its survival in the oral cavity and enables it to promote disease. Factors that promote A. actinomycetemcomitans colonization and persistence in the oral cavity include adhesins, bacteriocins, invasins and antibiotic resistance. It can interact with and adhere to all components of the oral cavity (the tooth surface, other oral bacteria, epithelial cells or the extracellular matrix). The adherence is mediated by a number of distinct adhesins that are elements of the cell surface (outer membrane proteins, vesicles, fimbriae or amorphous material). A. actinomycetemcomitans enhances its chance of colonization by producing actinobacillin, an antibiotic that is active against both streptococci and Actinomyces, primary colonizers of the tooth surface. The fact that A. actinomycetemcomitans resistance to tetracyclines, a drug often used in the treatment of periodontal disease, is on the rise is an added weapon. Periodontal pathogens or their pathogenic products must be able to pass through the epithelial cell barrier in order to reach and cause destruction to underlying tissues (the gingiva, cementum, periodontal ligament and alveolar bone). A. actinomycetemcomitans is able to elicit its own uptake into epithelial cells and its spread to adjacent cells by usurping normal epithelial cell function. A. actinomycetemcomitans may utilize these remarkable mechanisms for host cell infection and migration to deeper tissues. A. actinomycetemcomitans also orchestrates its own survival by elaborating factors that interfere with the host's defense system (such as factors that kill phagocytes and impair lymphocyte activity, inhibit phagocytosis and phagocyte chemotaxis or interfere with antibody production). Once the organisms are firmly established in the gingiva, the host responds to the bacterial onslaught, especially to the bacterial lipopolysaccharide, by a marked and continual inflammatory response, which results in the destruction of the periodontal tissues. A. actinomycetemcomitans has at least three individual factors that cause bone resorption (lipopolysaccharide, proteolysis-sensitive factor and GroEL), as well as a number of activities (collagenase, fibroblast cytotoxin, etc.) that elicit detrimental effects on connective tissue and the extracellular matrix. It is of considerable interest to know that A. actinomycetemcomitans possesses so many virulence factors but unfortunate that only a few have been extensively studied. If we hope to understand and eradicate this pathogen, it is critical that in-depth investigations into the biochemistry, genetic expression, regulation and mechanisms of action of these factors be initiated.

Abscess↗

PubMed1999

Identification of genes coding for exported proteins of Actinobacillus actinomycetemcomitans.

Random fusions of genomic DNA fragments to a partial gene encoding a signal sequence-deficient bacterial alkaline phosphatase were utilized to screen for exported proteins of Actinobacillus actinomycetemcomitans in Escherichia coli. Twenty-four PhoA(+) clones were isolated and sequenced. Membrane localization signals in the form of signal sequences were deduced from most of these sequences. Several of the deduced amino acid sequences were found to be homologous to known exported or membrane-associated proteins. The complete genes corresponding to two of these sequences were isolated from an A. actinomycetemcomitans lambda phage library. One gene was found to be homologous to the outer membrane lipoprotein LolB. The second gene product had homology with a Haemophilus influenzae protein and was localized to the inner membrane of A. actinomycetemcomitans.

Aggregatibacter actinomycetemcomitans↗

PubMed1998

Isolation and characterization of Fap1, a fimbriae-associated adhesin of Streptococcus parasanguis FW213.

An adhesin of Streptococcus parasanguis FW213, a primary colonizer of the tooth surface, has been purified from the culture medium by immunoaffinity chromatography. The purified protein has a molecular mass of 200 kDa and stains positively for carbohydrate. The amino-terminal sequence indicated that this protein represented a unique streptococcal surface protein. Immunogold labelling of the bacterium indicated that this protein was associated with fimbriae and designated Fap1 (fimbriae-associated protein). A polymerase chain reaction (PCR) product based on the amino terminus of Fap1 was used to probe an FW213 genomic library. A 9 kb fragment containing the fap1 gene was isolated and 2.5 kb have been sequenced. Generation of fap1 mutants by a single cross-over (Campbell insertion) or a non-polar allelic exchange abolished the expression of Fap1. The inactivation of fap1 resulted in a dramatic reduction in the expression of the long peritrichous fimbriae and adhesion to saliva-coated hydroxylapatite (SHA). Northern blots probed with an internal gene fragment of fap1 hybridized to a 9 kb transcript, which suggests that fap1 is transcribed as a polycistronic message. These data demonstrate that Fap1 is a unique streptococcal adhesin that is involved in the assembly of S. parasanguis FW213 fimbriae and adhesion to SHA.

Adhesins, Bacterial↗

PubMed1997

Models of invasion of enteric and periodontal pathogens into epithelial cells: a comparative analysis.

Bacterial invasion of epithelial cells is associated with the initiation of infection by many bacteria. To carry out this action, bacteria have developed remarkable processes and mechanisms that co-opt host cell function and stimulate their own uptake and adaptation to the environment of the host cell. Two general types of invasion processes have been observed. In one type, the pathogens (e.g., Salmonella and Yersinia spp.) remain in the vacuole in which they are internalized and replicate within the vacuole. In the other type, the organism (e.g., Actinobacillus actinomycetemcomitans, Shigella flexneri, and Listeria monocytogenes) is able to escape from the vacuole, replicate in the host cell cytoplasm, and spread to adjacent host cells. The much-studied enteropathogenic bacteria usurp primarily host cell microfilaments for entry. Those organisms which can escape from the vacuole do so by means of hemolytic factors and C type phospholipases. The cell-to-cell spread of these organisms is mediated by microfilaments. The investigation of invasion by periodontopathogens is in its infancy in comparison with that of the enteric pathogens. However, studies to date on two invasive periodontopathogens. A actinomycetemcomitans and Porphyromonas (Bacteroides) gingivalis, reveal that these bacteria have developed invasion strategies and mechanisms similar to those of the enteropathogens. Entry of A. actinomycetemcomitans is mediated by microfilaments, whereas entry of P. gingivalis is mediated by both microfilaments and microtubules. A. actinomycetemcomitans, like Shigella and Listeria, can escape from the vacuole and spread to adjacent cells. However, the spread of A. actinomycetemcomitans is linked to host cell microtubules, not microfilaments. The paradigms presented establish that bacteria which cause chronic infections, such as periodontitis, and bacteria which cause acute diseases, such as dysentery, have developed similar invasion strategies.

Actin Cytoskeleton↗

PubMed1997

Characterization of native and recombinant bone sialoprotein: delineation of the mineral-binding and cell adhesion domains and structural analysis of the RGD domain.

Bone sialoprotein is a small, sulfated, and phosphorylated integrin-binding glycoprotein apparently found only in tissues that eventually mineralize. Nondenatured bone sialoprotein (BSP) purified from rat osteosarcoma cell line (UMR 106-01 BSP) culture media is shown to have a hydroxyapatite Kd approximately 2.6 x 10(-9) M, perhaps the strongest affinity for this mineral of any of the matrix proteins. Both native BSP and a 47 kD fragment of UMR-BSP (Fragment 1 approximately 133A- approximately 265Y) are more potent inhibitors of seeded hydroxyapatite crystal growth than recombinant human BSP fragments lacking post-translational modifications. The recombinant proteins, however, do show reproducible inhibitory activity, suggesting that at least some of the strong mineral-binding properties are encoded directly within the protein sequence itself. BSP facilitates the adhesion of several cell types through its integrin binding (RGD) tripeptide sequence. Nuclear magnetic resonance (NMR) analysis of a 15N-enriched 59 amino acid recombinant domain containing the RGD tripeptide shows that the structure of this isolated domain is highly flexible with or without 5 mM calcium. Previous work has also shown that an endogenous fragment of UMR-BSP (Fragment 1) supports cell adhesion in the absence of the RGD sequence. In this report, non-RGD cell adhesion sites are localized within conserved amino- and carboxy-terminal tyrosine-rich domains of recombinant human BSP. Given the proximity of the latter non-RGD cell adhesion site to the RGD tripeptide, a model of BSP-receptor interactions is presented.

Amino Acid Sequence↗

PubMed1994

Chlorate-induced inhibition of tyrosine sulfation on bone sialoprotein synthesized by a rat osteoblast-like cell line (UMR 106-01 BSP).

Bone sialoprotein (BSP) is a major noncollagenous, RGD-containing glycoprotein found in the extracellular matrix of bone. The RGD sequence is flanked by two tyrosine-rich regions, which fit the established consensus requirements for tyrosine sulfation. Tyrosine sulfation is suggested to be important in the regulation of protein secretion and function. The role of this post-translational modification on the cell attachment activity and secretion of a highly sulfated form of BSP isolated from a rat osteoblast-like cell line (UMR 106-01 BSP) was investigated by inhibiting sulfation with chlorate. [35S]Sulfate, [3H]glucosamine, and [3H]tyrosine were used as metabolic precursors to monitor biosynthetic products. Chlorate was effective in inhibiting total [35S]sulfate incorporation by 90% without altering overall protein synthesis and secretion in cultures up to 72 h under serum-free conditions. Isolated proteoglycans and purified BSP were analyzed for sulfate incorporation. Proteoglycans isolated from the medium of cells treated with chlorate displayed a difference in the hydrodynamic properties of the molecules as compared with control cultures. An increase in the specific activity of proteoglycans labeled with [3H]glucosamine isolated from chlorate-treated cells was also observed suggesting a change in hexosamine metabolism induced by chlorate. BSP purified from the medium of chlorate-treated cells contained approximately 7% of the 35S incorporation as compared with nontreated control cultures. Quantification of sulfate incorporation into glycoconjugates versus tyrosine sulfate of BSP indicates that the amount of sulfate associated with N- and O-linked oligosaccharides was reduced by approximately 97%, while that on tyrosine residues was reduced by approximately 90%. Using normal human bone cells, the cell attachment activity of the reduced sulfate form of BSP was nearly equivalent to that of the fully sulfated product.

Animals↗

PubMed1994

Identification of an immunoglobulin Fc receptor of Actinobacillus actinomycetemcomitans.

Actinobacillus actinomycetemcomitans expresses proteins that bind to the Fc portion of immunoglobulins. The immunoglobulin Fc receptors on the surface of A. actinomycetemcomitans were detected by the binding of biotinylated human or murine Fc molecules to strain SUNY 465 adsorbed to the bottom of microtiter wells. Biotinylated Fc binding was inhibited by unlabeled Fc molecules and human plasma. Fc receptors were identified by the binding of biotinylated Fc molecules to bacterial membrane proteins separated by polyacrylamide gel electrophoresis and transferred to nitrocellulose. Multiple bands were identified, and the major Fc-binding protein was determined to be a heat-modifiable protein. This protein migrated with approximate molecular weights of 25,000 and 32,000 (unheated and heated, respectively). Amino-terminal sequence analysis of this protein revealed a sequence identical to the heat-modifiable protein described for A. actinomycetemcomitans ATCC 43718. This protein sequence exhibits significant homology with the N termini of outer membrane protein A (OmpA) of Escherichia coli and related OmpA-like proteins from other gram-negative bacteria.

Aggregatibacter actinomycetemcomitans↗

PubMed1994

Adhesion of Actinobacillus actinomycetemcomitans to a human oral cell line.

Two quantitative, rapid assays were developed to study the adhesion of Actinobacillus actinomycetemcomitans, an oral bacterium associated with periodontal disease, to human epithelial cells. The human oral carcinoma cell line KB was grown in microtiter plates, and adherent bacteria were detected by an enzyme-linked immunosorbent assay with purified anti-A. actinomycetemcomitans serum and horseradish peroxidase-conjugated secondary antibody or [3H]thymidine-labeled bacteria. Adhesion was found to be time dependent and increased linearly with increasing numbers of bacteria added. Variation in the level of adhesion was noted among strains of A. actinomycetemcomitans. Adhesion was not significantly altered by changes in pH (from pH 5 to 9) but was sensitive to sodium chloride concentrations greater than 0.15 M. Pooled human saliva was inhibitory for adhesion when bacteria were pretreated with saliva before being added to the cells. Pretreatment of the KB cells with saliva did not inhibit adhesion. Protease treatment of A. actinomycetemcomitans reduced adhesion of the bacteria to KB cells. The data are consistent with the hypothesis that a protein(s) is required for bacterial adhesion and that host components may play a role in modulating adhesion to epithelial cells.

Aggregatibacter actinomycetemcomitans↗

PubMed1994

Interactions of matrix proteins from mineralized tissues with octacalcium phosphate.

Acidic matrix macromolecules, present in many mineralized tissues, including those of vertebrates, are thought to be involved in controlling crystal formation. Little, however, is known about their in vivo functions, particularly in relation to calcium-phosphate-containing crystals. The manner in which a variety of synthetic and natural acidic macromolecules interact in vitro with crystals of octacalcium phosphate (OCP) has been studied. Interactions were assessed by examining changes in morphology of the crystals resulting from preferential interaction of the additive with some crystal faces and not others. Macromolecules rich in acidic amino acids, with or without polysaccharides, such as polyaspartate and mollusk shell proteins respectively, were shown to interact preferentially with rows of Ca ions exposed on the hydrated plate surface of OCP crystals. In contrast, the phosphorylated proteins, phosphophoryn and phosvitin, interacted specifically with the apatite-like motifs on the OCP side faces. BSP did not interact specifically with OCP, under the experimental conditions used. The observation that these classes of acidic macromolecules recognize different crystal faces should be taken into account when evaluating functions of acidic matrix macromolecules in mineralized tissues.

Animals↗

PubMed1993

Purification and fragmentation of nondenatured bone sialoprotein: evidence for a cryptic, RGD-resistant cell attachment domain.

Bone sialoprotein (BSP), a small (approximately 80,000 M(r)) integrin binding, RGD-containing bone matrix glycoprotein, has been purified in milligram quantities from the serum-free medium of the rat osteosarcoma cell line UMR-106-BSP using nondenaturing conditions. Routine protein purification without serine protease inhibitors or reducing agents consistently resulted in three major fragments. The largest fragment (E1) started at amino acid 117 and did not bind to antibodies made to the RGD region of the protein. Furthermore, the smallest fragment (E3), was shown by sequencing to contain the RGD region of the protein. Digestion of intact BSP with highly purified chymotrypsin also resulted in a large fragment (C1) with properties nearly identical to those of E1. The large, non-RGD-containing fragments, E1 and C1, as well as the intact BSP, supported attachment by normal human bone cells and human skin fibroblasts in vitro. Attachment to the intact BSP was totally blocked by 0.4 mM GRGDS peptide. Both preparations of skin fibroblasts and approximately half of the preparations of normal human bone cells, however, also would not attach to the E1 and C1 fragments in the presence of 0.4 mM GRGDS peptide. In contrast, half of the bone cell preparations had significant attachment activity to E1 (> 50%) and C1 (> 25%) in the presence of 0.4 mM GRGDS peptide. These data suggest that cleavage of the BSP results in either (1) the exposure of a previously unavailable or cryptic cell attachment site or (2) a conformational change that increases the affinity of the complex between a non-RGD-encoded binding region of the E1 and C1 fragments and at least one receptor. The possible homology of the second, non-RGD-suppressible site of BSP with the second cell attachment site on the gamma chain of fibrinogen is discussed.

Adult↗

PubMed1993

Structure and molecular regulation of bone matrix proteins.

The organic matrix of bone contains several protein families, including collagens, proteoglycans, and glycoproteins, all of which may be extensively modified by posttranslational events, such as phosphorylation and sulfation. Many of the glycoproteins contain Arg-Gly-Asp (RGD), the integrin-binding sequence, within their structure, whereas other constituent proteins contain gamma-carboxyglutamic acid. The deposition of bone matrix by cells in the osteoblastic lineage is regulated by extrinsic factors, such as systemic and local growth factors and physical forces, and factors that are intrinsic to the cell, such as position in the cell cycle, maturational stage, and developmental age of the donor. Recent studies of several bone matrix gene promoters have identified cis- and trans-acting elements that are responsible for gene activity, although the precise sequence of regulatory events is not known. Development of in vitro assays, coupled with studies of the appearance of these proteins during development in vivo, provides insight into the functions of these proteins during the various stages of bone metabolism. Potential roles for these proteins include proliferation and maturation of stem cells, formation of matrix scaffolding elaborated by bone-forming cells, modeling, and remodeling. Changes in the functional properties of the extracellular matrix may be involved in a variety of disease processes, including osteoporosis and oral bone loss.

Bone Matrix↗

PubMed1991

Osteogenesis imperfecta: changes in noncollagenous proteins in bone.

The noncollagenous proteins osteonectin, bone sialoprotein, osteocalcin, the small proteoglycan decorin (PG II), and alpha 2-HS glycoprotein (which is synthesized in the liver but highly concentrated in bone) were measured in extracts of cortical bone from 3 type I, 2 type II, 8 type III and 13 type IV patients with osteogenesis imperfecta (OI) and from 7 control subjects. Osteonectin was found to be reduced in the bone of all OI patients. The bone from severely affected type III OI patients contained the lowest levels of osteonectin. In contrast, bone sialoprotein was found to be elevated in the bones of OI patients. The highest levels were found in individuals classified as type IV patients. Osteocalcin and alpha 2-HS glycoprotein concentrations were increased in all OI patients. Decorin levels were not significantly altered in OI bones compared to controls. These changes in the concentrations of the noncollagenous proteins may contribute to the fragility of the OI bone by interfering with complete mineralization and/or normal tissue architecture.

Adolescent↗

PubMed1990

Detection of procollagen biosynthesis using peptide-specific antibodies.

Peptides corresponding to selected sequences of the alpha 1 chain of the COOH propeptide of type I and type III human procollagen were synthesized and used as antigens to develop polyclonal and monoclonal antibodies. The antibodies were shown to be epitope specific using a peptide-based solid phase enzyme-linked immunoadsorbent assay. The antibodies were specific for the appropriate procollagens and the COOH propeptides isolated from serum-free culture supernatants of human skin fibroblasts. The rabbit antisera directed to the type I synthetic peptide bound the intact procollagen molecule and both the procollagen alpha 1(I) and alpha 2(I) chains after the reduction of the disulfide bonds. In addition, the antisera bound intact type I COOH propeptide, generated by bacterial collagenase treatment of procollagen, and the individual chains of the propeptide after reduction. In contrast, a monoclonal antibody to the type I peptide was able to bind only to the reduced form of the COOH propeptide. Both rabbit polyclonal and murine monoclonal antibodies directed to the type III synthetic peptide bound the intact and the individual chains of type III procollagen as well as the intact and reduced forms of the type III COOH propeptide. The antibodies have been used to detect procollagen synthesis in two human osteosarcoma cell lines and the differential expression of procollagen in the culture medium of rat lung fibroblasts grown in the presence or absence of glucocorticoids.

Amino Acid Sequence↗

PubMed1986

Glucocorticoids decrease the synthesis of type I procollagen mRNAs.

Glucocorticoids selectively decrease procollagen synthesis in animal and human skin fibroblasts. beta-Actin content and beta-actin mRNA are not affected by glucocorticoid treatment of chick skin fibroblasts. The inhibitory effect of glucocorticoids on procollagen synthesis is associated with a decrease in total cellular type I procollagen mRNAs in chick skin fibroblasts. These effects of dexamethasone are receptor mediated as determined by pretreatment with the glucocorticoid antagonists progesterone and RU-486 and with the agonist beta-dihydrocortisol. Dexamethasone has a small but significant inhibitory effect on cell growth of chick skin fibroblasts. The ability of this corticosteroid to decrease the steady-state levels of type I procollagen mRNAs in nuclei, cytoplasm, and polysomes varies. The largest decrease of type I procollagen mRNAs is observed in the nuclear and cytoplasmic subcellular fractions 24 h after dexamethasone treatment. Type I procollagen hnRNAs are also decreased as determined by Northern blot analysis of total nuclear RNA. The synthesis of total cellular type I procollagen mRNAs is reversibly decreased by dexamethasone treatment. In addition the synthesis of total nuclear type I procollagen mRNA sequences is decreased at 2, 4, and 24 h following the addition of radioactive nucleoside and dexamethasone to cell cultures. Although the synthesis of pro alpha 1(I) and pro alpha 2(I) mRNAs is decreased in dexamethasone-treated chick skin fibroblasts, the degradation of the total cellular procollagen mRNAs is not altered while the degradation of total cellular RNA is stabilized. These data indicate that the dexamethasone-mediated decrease of procollagen synthesis in embryonic chick skin fibroblasts results from the regulation of procollagen gene expression.

Actins↗

PubMed1985

Human acetylcholinesterase. Immunochemical studies with monoclonal antibodies.

Monoclonal antibodies were used to investigate the immunochemistry of human erythrocyte acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7). A series of experiments on the sedimentation velocity and Stokes radius of acetylcholinesterase and its immune complexes indicated that each antibody recognized a single high-affinity binding site (epitope) on the monomeric enzyme. Further analysis suggested that the antibody-binding sites were replicated on multimeric enzyme forms but were subject to steric hindrance between nearby IgG molecules or adjacent enzyme subunits. The cellular localization of the epitopes was studied by measuring the binding of monoclonal antibodies to the cholinesterase of intact erythrocytes. The results implied that most of the epitopes are exposed to the external media. However, one antibody failed to bind to intact cells, despite a relatively high affinity for detergent-solubilized antigen, possibly because its epitope is buried in the lipid bilayer.

Acetylcholinesterase↗

PubMed1985

Monoclonal antibodies to rabbit brain acetylcholinesterase: selective enzyme inhibition, differential affinity for enzyme forms, and cross-reactivity with other mammalian cholinesterases.

Eleven unique monoclonal IgG antibodies were raised against rabbit brain acetylcholinesterase (AChE, EC 3.1.1.7), purified to electrophoretic homogeneity by a two-step procedure involving immunoaffinity chromatography. The apparent dissociation constants of these antibodies for rabbit AChE ranged from about 10 nM to more than 100 nM (assuming one binding site per catalytic subunit). Species cross-reactivity was investigated with crude brain extracts from rabbit, rat, mouse cat, guinea pig, and human. One antibody bound rabbit AChE exclusively; most bound AChE from three or four species; two bound enzyme from all species tested. Identical, moderate affinity for rat and mouse brain AChE was displayed by two antibodies; two others were able to distinguish between these similar antigens. Nine of the antibodies had lowered affinity for AChE in the presence of 1 M NaCl, but two were salt resistant. Analysis of mutual interferences in AChE binding suggested that certain of the antibodies were competing for nearby epitopes on the AChE surface. One antibody was a potent AChE inhibitor (IC50 = 10(-8) M), blocking up to 90% of the enzyme activity. Most of the antibodies were less able to bind the readily soluble AChE of detergent-free brain extracts than the AChE which required detergent for solubilization. The extreme case, an antibody that was unable to recognize nearly half of the "soluble" AChE, was suspected of lacking affinity for the hydrophilic enzyme form.

Acetylcholinesterase↗