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Biomedical subjects

L D Kohn

Publications and source records attributed to L D Kohn.

At least 271 records · Page 15Linked to original sources
PubMed1974

Calf tendon procollagen peptidase: its purification and endopeptidase mode of action.

The procollagen peptidase activity of calf tendon has been purified. The enzyme has a high degree of specificity for native procollagen and converts both pro alpha1 and alpha2, to alpha1 and alpha2, respectively. The purified enzyme is an endopeptidase which excises the amino terminal peptide extensions of the precursor chains in block; the molecular size and amino-acid composition of the excised peptides compare favorably with those predicted in previous reports. Antisera to the enzyme and to procollagen have been prepared and have been used to characterize the enzyme, the enzymatically excised peptides, and the enzyme-peptide complex in reaction mixtures.

Amino Acids↗
PubMed1974

D-lactate dehydrogenase binding in Escherichia coli dld- membrane vesicles reconstituted for active transport.

When membrane vesicles prepared from a D-lactate dehydrogenase mutant of E. coli ML 308-225 are treated with a homogeneous preparation of D-lactate dehydrogenase, the enzyme binds to the vesicles and they regain the capacity to catalyze D-lactate oxidation and D-lactate-dependent active transport. Although membranebound enzyme increases linearly with addition of increasing quantities of enzyme, reconstituted transport activity and D-lactate oxidation are saturable functions of the amount of enzyme bound. The maximal specific transport activity obtained in the reconstituted system is similar in magnitude to that of wild type vesicles. Titration studies with 2-(N-dansyl)-aminoethyl-beta-D-thiogalactoside demonstrate that there is at least a 7- to 8-fold excess of lac carrier protein relative to D-lactate dehydrogenase. Hydroxybutynoate-inactivated enzyme does not bind to the vesicles, indicating that the coenzyme moiety is critically involved in binding. Conformational changes are also apparently involved since 0.6 M guanidine.HCl is required for optimal binding and reconstitution. The relative unreactivity of reconstituted vesicles towards vinylglycolic acid suggests that D-lactate dehydrogenase is bound to the outer surface of the reconstituted vesicles.

Binding Sites↗
PubMed1974

The contribution of subunits of thyroid stimulating hormone to the binding and biological activity of thyrotropin.

The binding of bovine TSH (thyroid stimulating hormone), LH (luteinizing hormone), and their subunits to the TSH receptor of beef thyroid membranes was compared to stimulation by these agents of adenylate cyclase [ATP pyrophosphate-lyase(cyelizing), EC 4.6.1.1] in the same membranes, glucose oxidation in dog thyroid slices, and the secretory process in mouse thyroids in vitro (colloid droplet formation) and in vivo (hormone release). The beta-subunits of TSH and LH can bind to the TSH receptor and can activate thyroid function in vitro. In contrast, the alpha-subunit of TSH binds negligibly to the TSH receptor and has very low potency for stimulation of thyroid function (except for colloid droplet formation). Neither binding nor the biological activity of the beta-subunits can be accounted for by TSH contamination, whereas this cannot be ruled out for alpha-TSH. LH binds to the TSH receptor even better than the beta-subunit of TSH but the increased binding does not result in a corresponding activation of thyroid function. Neither alpha- nor beta-TSH alone can induce more than 4-8% of the response to intact TSH in any of the investigated parameters. It is proposed that the beta-subunit has within its structure the primary determinants which are necessary to stimulate biological activity, whereas the alpha-subunit imposes conformational changes on the beta-subunit which in intact TSH promote binding and biological activity commensurately but in LH promote only binding.

Adenylyl Cyclases↗
PubMed1973

Defect in conversion of procollagen to collagen in a form of Ehlers-Danlos syndrome.

Three patients with a form of the Ehlers-Danlos syndrome, a generalized disorder of connective tissue, have detectable amounts of procollagen in extracts of their skin and tendon. The activity of procollagen peptidase, the enzyme that converts procollagen to collagen, is reduced in cultures of fibroblasts. The clinical manifestations of this syndrome may be related to impaired enzymatic conversion of procollagen to collagen. Cultures of skin fibroblasts from these patients have an increased rate of synthesis of collagenous protein (collagen and procollagen), possibly related to the inability of these cells to convert procollagen to collagen.

Carbon Radioisotopes↗
PubMed1973

Cell-free synthesis of procollagen: L-929 fibroblasts as a cellular model for dermatosparaxis.

A cell-free system that actively synthesizes collagen was prepared from L-929 fibroblasts. Chromatographic and electrophoretic techniques were used to demonstrate that the only collagenous products are pro alpha(1) and pro alpha(2) chains. The collagen synthesized by the cell-free system was also compared to the collagen extracted from the cells. The cellular collagen was composed of aggregates of pro-alpha chains, while no alpha chains were found. Procollagen peptidase activity could not be detected in the cells, and the activity present in the medium was low, comparable to that in dermatosparaxic cell cultures. These properties indicate that L-929 cells may be a model system for dermatosparaxis.

Animals↗
PubMed1972

The binding of ( 3 H)thyrotropin and an 3 H-labeled exophthalmogenic factor by plasma membranes of retro-orbital tissue.

Plasma membranes of cells from retroorbital tissue have been prepared from the Harderian glands of guinea pigs and have been characterized as being reasonably free of other subcellular structures by electron microscopy and by enzyme-marker analyses. Both bovine thyrotropin and a proteolytic derivative of bovine thyrotropin with exophthalmogenic activity but without thyroid-stimulating activity specifically bind to these membranes. Gammaglobulin from the sera of patients with malignant exophthalmos increases the binding of both pituitary factors, whereas binding is not similarly increased by gammaglobulin from the sera of individuals who are not exophthalmic. The increased binding caused by the gammaglobulin from exophthalmic patients is the same whether the sera are positive or negative for the long-acting thyroid stimulator. Present binding experiments do not indicate a direct interaction between the gammaglobulin and the plasma membranes of the cells from Harderian glands. A mechanism for the pathogenesis of human exophthalmos is proposed on the basis of these data.

Animals↗
PubMed1972

In vitro synthesis of procollagen on polysomes.

The major collagenous products synthesized in a cell-free polysome preparation are pro-alpha1 and pro-alpha2, formed in a ratio of 2:1. They are the precursor forms of alpha1 and alpha2 chains of normal connective-tissue collagen that are also formed in small amounts. A procollagen peptidase activity has been demonstrated in the supernatant fraction that can account for the formation of alpha1 and alpha2 chains from their precursors. The polysomal system is activated by a salt extract of reticulocyte ribosomes and is inhibited by aurintricarboxylic acid, suggesting that the polysomes are able to initiate protein synthesis.

Animals↗
PubMed1971

Procollagen peptidase: an enzyme excising the coordination peptides of procollagen.

A heritable connective tissue disorder of cattle, dermatosparaxis, is characterized by an extreme fragility of the skin and the presence of additional peptides at the N-terminal extremities of the collagen alpha chains, p-alpha(1) and p-alpha(2). The existence of an enzyme activity is demonstrated in normal connective tissues that is capable of cleaving these additional N-terminal peptides from dermatosparaxic collagen. The activity is demonstratable with dermatosparaxic collagen in solution, as well as with reconstituted dermatosparaxic collagen fibrils polymerized in vitro. It has a pH optimum of about 7.0 and is inhibited by EDTA and mercaptoethanol. Differences in K(m) and V(max) values exist depending on the substrate utilized, i.e., p-alpha(1) or p-alpha(2); and the presence of additional amounts of one substrate, p-alpha(1), alters the concentration requirement for the second substrate, p-alpha(2). The product of the excision reaction with p-alpha(1) as substrate is an equimolar amount of normal alpha(1) monomer; the product when p-alpha(2) is substrate is an equimolar amount of normal alpha(2) monomer. The enzyme is present in normal calf skin, tendon, aorta, cartilage, and lung; it can be demonstrated in the skin of rats and humans. The enzyme activity is absent in dermatosparaxic connective tissues, thus suggesting that dermatosparaxis is caused by the absence of a normal enzyme function rather than by the production of an abnormal collagen.

Animals↗