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S S Carlson

Publications and source records attributed to S S Carlson.

11 recordsLinked to original sources
PubMed1991

A large chondroitin sulfate basement membrane-associated proteoglycan exists as a disulfide-stabilized complex of several proteins.

Proteoglycan (PG)-1000 (formerly TAP-1) is a large (Mr = 10(6)) highly glycosylated chondroitin sulfate proteoglycan found associated with Schwann cell and electrocyte basement membranes in elasmobranch electric fish. Previously, purified PG-1000 was visualized in the electron microscope as a "bottlebrush" structure about 345 nm long with about 20 side projections of 113 nm. This molecule was characterized with material purified from electric organ under denaturing and reducing conditions. Here we report that PG-1000, when purified under denaturing conditions without exposure to a reducing agent, exists as a complex of several proteins. In addition to PG-1000, this complex consists of a somewhat smaller, heavily glycosylated protein (beta component) and three smaller proteins with Mr values of 39,000, 21,000, and 18,000. The complex remains intact when exposed to denaturing and non-reducing conditions but falls apart in denaturing and reducing conditions. Presumably the complex is stabilized by disulfide bonds. The beta component of the PG-1000 complex is probably a proteoglycan. However, unlike PG-1000, the beta component does not contain chondroitin sulfate chains and lacks the epitope, T1, that is found on PG-1000. Both molecules share a protease-insensitive antigenic site, SV4, which is probably a modified keratan sulfate epitope. Evidence for the identity of this antigen is that it is found as a minor subfraction in commercial preparations of shark cartilage chondroitin and corneal keratan sulfates but not in other glycosaminoglycan preparations. These SV4 antigens are resistant to chondroitin ABC lyase digestion. However, the SV4 antigen in commercial keratan sulfate is cleaved by keratinase to a smaller antigenic fragment.

Animals↗
PubMed1987

Nerve terminal anchorage protein 1 (TAP-1) is a chondroitin sulfate proteoglycan: biochemical and electron microscopic characterization.

The plasma membranes of the nerve terminal and the postsynaptic cell of electric organ are separated by a basal lamina. We have purified, biochemically characterized, and visualized in the electron microscope a macromolecule which appears to anchor the nerve terminal to this basal lamina. This molecule, terminal anchorage protein 1 (TAP-1) is associated with the nerve terminal membrane of electric organ, has the properties of an integral membrane protein, and is tightly bound to the extracellular matrix (Carlson, S.S., P. Caroni, and R.B. Kelly. 1986. J. Cell Biol. 103:509-520). TAP-1 can be solubilized from an electric organ extracellular matrix preparation with guanidine-HCl/3-[(3-cholamidopropyl)-dimethylammnio]-1-propane sulfonate and purified by a combination of permeation chromatography on Sephacryl S-1000, sedimentation velocity, and ion exchange chromatography on DEAE Sephacel. The total purification from electric organ is 91-fold and results in at least 86% purity. Digestion of the molecule with chondroitin ABC or AC lyase produces a large but similar shift in the molecular weight of the molecule on SDS-PAGE. The presence of chondroitin-4- or 6-sulfate is confirmed by identification of the isolated glycosaminoglycans with cellulose acetate electrophoresis. Gel filtration of the isolated chains indicates an average molecular weight of 42,000. Digestion of TAP-1 with other glycosaminoglycan lyases such as heparitinase indicates that only chondroitin sulfate is present. These results demonstrate that TAP-1 is a proteoglycan. Visualization of TAP-1 in the electron microscope reveals a "bottlebrush" structure expected for a proteoglycan. The molecule has an average total length of 345 +/- 17 nm with 20 +/- 2 side projections of 113 +/- 5 nm in length. These side projections are presumably the glycosaminoglycan side chains. From this structure, we predict that the TAP-1 glycosaminoglycan side chains should have a molecular weight of approximately 50,000, which is in close agreement with the biochemical studies. Both biochemical and morphologic data indicate that TAP-1 has a relative molecular weight of approximately 1.2 X 10(6). The large size of TAP-1 suggests that this molecule could span the synaptic cleft and make a significant contribution to the structure of the nerve terminal basal lamina of electric organ.

Animals↗
PubMed1986

A nerve terminal anchorage protein from electric organ.

The nerve terminal and the postsynaptic receptor-containing membranes of the electric organ are both linked to the basal lamina that runs between them. We have identified an extracellular matrix protein whose physical properties suggest it anchors the nerve terminal to the basal lamina. The protein was identified because it shares an epitope with a proteoglycan component of electric organ synaptic vesicles. It too behaves like a proteoglycan. It is solubilized with difficulty from extracellular matrix fractions, elutes from DEAE Sephacel at pH 4.9 only at high ionic strength, and binds to a laminin affinity column from which it can be eluted with heparin. Under denaturing conditions it sediments rapidly and has a large excluded volume although it can be included in Sephacryl S-1000 columns. This large, highly charged extracellular matrix molecule can be readily reconstituted into liposomes consistent with the presence of a hydrophobic tail. By immunoelectron microscopy the antigen is found both in synaptic vesicles and on the plasma membrane of the nerve terminal. Since this is the first protein described that links the nerve terminal membrane to the extracellular matrix, we propose calling it terminal anchorage protein one (TAP-1).

Animals↗
PubMed1985

Localization and axonal transport of immunoreactive cholinergic organelles in rat motor neurons--an immunofluorescent study.

Antisera produced in rabbits against pure fractions of cholinergic vesicles from Narcine brasiliensis were used to study cholinergic organelles in rat motor neurons. The indirect immunofluorescence method was used on perfusion-fixed material. The rats were surgically sympathectomized to remove sympathetic adrenergic and cholinergic nerves from the sciatic nerve. In the intact animal immunoreactive material, likely to represent cholinergic vesicles, was observed in motor endplates, identified by labelling with rhodamine-conjugated alpha-bungarotoxin or with subsequent acetylcholinesterase staining. The motor perikarya contained very little immunoreactive material. Non-terminal axons were virtually devoid of immunofluorescence in the intact animal. After crushing the sciatic nerve, immunoreactive material (likely to represent axonal cholinergic organelles) accumulated rapidly on both sides of the crush, indicating a rapid bidirectional transport. The transport was sensitive to local application of mitotic inhibitors. The axons which accumulated immunoreactive organelles were motor axons, as demonstrated by various procedures: Cutting of ventral roots prevented accumulation of immunoreactive material in the nerve. Deafferentation did not notably influence accumulations of immunoreactive material. Ligated axons with immunoreactive material were acetylcholinesterase positive when identification was made on the same section; the intra-axonal distribution of immunoreactive material and acetylcholinesterase was not identical, however, and the Narcine antisera did not cross-react with bovine acetylcholinesterase in a solid phase immunoassay. Most axons in ventral roots, but not in dorsal roots, accumulated strongly fluorescent immunoreactive material, while axons in dorsal roots contained weakly fluorescent material. On the other hand, substance P-like immune reactivity was present in many dorsal root axons, but only very rarely in ventral roots. It is suggested that the antisera against Narcine cholinergic vesicles can be used as a marker for cholinergic organelles in the motor neuron, and may be an important tool for studying the axonal cholinergic vesicles. It cannot, however, be used to identify cholinergic structures in unknown locations because it recognizes common antigenic determinants in transmitter organelles of other nerves, e.g. adrenergic nerves. The axonal cholinergic organelles may carry important molecules, other than acetylcholine to the nerve endings.

Animals↗
PubMed1978

Purification of synaptic vesicles from elasmobranch electric organ and the use of biophysical criteria to demonstrate purity.

We have purified cholinergic synaptic vesicles from the electric organs of two related marine elasmobranchs, Torpedo californica and Narcine brasiliensis, to a specific activity higher than had previously been obtained. We have demonstrated the homogeneity of the vesicles by biophysical criteria. The purification scheme consisted of differential centrifugation, flotation equilibrium in sucrose density gradients, and permeation chromatography on glass bead columns of average pore size 3000 A. Our criteria for purity were that bound acetylcholine, bound nucleotide triphosphate, protein, and lipid--phosphorus behave identically when vesicles were analyzed by procedures which depend on vesicle size, density, and charge. Contaminants were not detected when vesicles were fractionated by preparative and analytical sedimentation, by preparative equilibrium sedimentation using glycerol density gradients, or by electrophoresis in Ficoll density gradients. Pure synaptic vesicles, which have been purified 290-fold from the initial homogenate, contain per mg of protein: 8 mumol of acetylcholine, 3 mumol of ATP, and 7 mumol of lipid phosphorus. These procedures may be of general value in the purification of membrane vesicles.

Animals↗
PubMed1977

Primary structure of mouse, rat, and guinea pig cytochrome c.

For immunochemical and evolutionary reasons we determined the primary structure of cytochrome c from two strains of laboratory mice. Thioacetylthioethane and thioacetylthioglycolic acid were used in addition to conventional reagents for sequence determinations. The sequence was found to be identical with that of the rabbit except for residues 44 and 89 and consistent with the peptide compositional data reported by Hennig (Hennig, B. (1975), Eur. J. Biochem. 55, 167-183). The rat cytochrome c cymotryptic peptides were identical with those of the mouse in amino acid composition and amino-terminal residues. Further, peptide maps of cytochromes c of the guinea pig and two strains of rat indicate that all these animals have the same cytochrome c as the laboratory mouse. It is concluded that rodent cytochromes c are evolutionarily conservative and that there is no evidence for a generation-time effect in cytochrome c evolution.

Amino Acid Sequence↗
PubMedDate not supplied

Presynaptic neurones may contribute a unique glycoprotein to the extracellular matrix at the synapse.

As the extracellular matrix at the original site of a neuromuscular junction seems to play a major part in the specificity of synaptic regeneration, considerable attention has been paid to unique molecules localized to this region. Here we describe an extracellular matrix glycoprotein of the elasmobranch electric organ that is localized near the nerve endings. By immunological criteria, it is synthesized in the cell bodies, transported down the axons and is related to a glycoprotein in the synaptic vesicles of the neurones that innervate the electric organ. It is apparently specific for these neurones, as it cannot be detected elsewhere in the nervous system of the fish. Therefore, neurones seem to contribute unique extracellular matrix glycoproteins to the synaptic region. Synaptic vesicles could be involved in transporting these glycoproteins to or from the nerve terminal surface.

Animals↗