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Carl-Wilhelm Vogel

Publications and source records attributed to Carl-Wilhelm Vogel.

6 recordsLinked to original sources
PubMed2006

Actin-binding protein filamin A is displayed on the surface of human neuroblastoma cells.

We previously reported the identification of natural human IgM antibodies, which recognize a M(r) 260 000 surface protein (NB-p260) and induce both complement-mediated cytotoxicity and apoptosis of human neuroblastoma cells. NB-p260 was shown to belong to the family of filamin proteins. Filamin A is a high molecular weight actin-binding protein, previously thought to be only located intracellularly. Here we show that NB cells as well as three NB-unrelated human cell lines express filamin A also on the cell surface. Our findings suggest new biological functions for filamins, including a role as mediators in anti-NB IgM-induced apoptosis, and they add to the growing body of evidence of the interaction of cytoskeletal proteins with the extracellular matrix.

Actins↗
PubMed2004

Structure and function of recombinant cobra venom factor.

Cobra venom factor (CVF) is the complement-activating protein from cobra venom. It is a structural and functional analog of complement component C3. CVF functionally resembles C3b, the activated form of C3. Like C3b, CVF binds factor B, which is subsequently cleaved by factor D to form the bimolecular complex CVF,Bb. CVF,Bb is a C3/C5 convertase that cleaves both complement components C3 and C5. CVF is a three-chain protein that structurally resembles the C3b degradation product C3c, which is unable to form a C3/C5 convertase. Both C3 and CVF are synthesized as single-chain prepro-proteins. This study reports the recombinant expression of pro-CVF in two insect cell expression systems (baculovirus-infected Sf9 Spodoptera frugiperda cells and stably transfected S2 Drosophila melanogaster cells). In both expression systems pro-CVF is synthesized initially as a single-chain pro-CVF molecule that is subsequently proteolytically processed into a two-chain form of pro-CVF that structurally resembles C3. The C3-like form of pro-CVF can be further proteolytically processed into another two-chain form of pro-CVF that structurally resembles C3b. Unexpectedly, all three forms of pro-CVF exhibit functional activity of mature, natural CVF. Recombinant pro-CVF supports the activation of factor B in the presence of factor D and Mg2+ and depletes serum complement activity like natural CVF. The bimolecular convertase pro-CVF,Bb exhibits both C3 cleaving and C5 cleaving activity. The activity of pro-CVF and the resulting C3/C5 convertase is indistinguishable from CVF and the CVF,Bb convertase. The ability to produce active forms of pro-CVF recombinantly ensures the continued availability of an important research reagent for complement depletion because cobra venom as the source for natural CVF will be increasingly difficult to obtain as the Indian cobra is on the list of endangered species. Experimental systems to express pro-CVF recombinantly will also be invaluable for studies to delineate the structure and function relationship of CVF and its differences from C3 as well as to generate human C3 derivatives with CVF-like function for therapeutic complement depletion ("humanized CVF").

Animals↗
PubMed2004

Recombinant cobra venom factor.

Cobra venom factor (CVF) is the complement-activating protein from cobra venom. CVF is a three-chain protein that functionally resembles C3b, the activated form of complement component C3. Like C3b, CVF forms a C3/C5 convertase with factor B in the presence of factor D and Mg(2+). Although CVF exhibits functional activity of C3b, it structurally resembles the C3b degradation product C3c, which is not able to form a C3/C5 convertase. CVF has become an important research tool to decomplement laboratory animals in order to study the role of complement in host defense, immune response, and pathogenesis of disease. As the Asian cobras of the Naja species are on the list of endangered species, cobra venom as the source for CVF has become increasingly difficult to obtain. Methods have been developed to recombinantly produce active forms of CVF. This manuscript reviews the production of recombinant pro-CVF using both prokaryotic and eukaryotic expression systems. The recombinant production of pro-CVF in two insect cell expression systems (baculovirus-infected Sf9 Spodoptera frugiperda cells, stably transfected S2 Drosophila melanogaster cells) generates three forms of pro-CVF: single-chain pro-CVF resembling pro-C3, a two-chain form of pro-CVF resembling C3, and another two-chain form of pro-CVF resembling C3b. All three forms of pro-CVF exhibit functional activity of mature, natural CVF. Recombinant pro-CVF supports the activation of factor B in the presence of factor D and Mg(2+), forms a bimolecular convertase pro-CVF,Bb that exhibits cleaving activity for both C3 and C5, and depletes the serum complement activity. The activity of pro-CVF and the resulting C3/C5 convertase is indistinguishable from CVF and the CVF,Bb convertase. Recombinant production of functionally active forms of pro-CVF ensures the availability of an important research reagent for future research involving complement depletion. The experimental systems to recombinantly produce active forms of CVF will also be invaluable for studies to delineate the structure/function relationship of CVF and its differences from C3, and to generate human C3 derivatives with CVF-like function ("humanized CVF") for therapeutic complement depletion.

Animals↗
PubMed2004

Preparation of immunoconjugates using antibody oligosaccharide moieties.

Heterobifunctional crosslinking reagents are small molecular weight chemicals containing two different reactive groups that have become important tools in generating conjugates of two different biomolecules, such as two proteins. The resulting bioconjugates are hybrid molecules or proteins, a new category of biomolecules that exhibit the combined functions of the two parent biomolecules. An important category of hybrid proteins are conjugates of antibodies with other effector molecules, such as drugs or toxins. These antibody conjugates or immunoconjugates have a variety of the applications in medicine, with particular emphasis on the treatment of cancer. The most commonly used heterobifunctional crosslinking reagents for the synthesis of antibody conjugates contain an N-hydroxysuccinimide ester moiety, which allows derivatization of amino groups in proteins. The chemical modification of a functionally important amino group in the antigen-binding region of an antibody causes impairment or loss of the antigen binding function, resulting in a defective antibody conjugate that lacks one of its component functions. Furthermore, even if the chemical derivatization does not affect the antigen binding function, the subsequent coupling of an effector protein at or near the antigen-binding region can also cause the loss of the antigen binding function for steric reasons. In this chapter, heterobifunctional crosslinking reagents are described that allow the generation of antibody conjugates where the effector proteins are coupled to the antibody carbohydrate moieties. Because antibody carbohydrate moieties are distal from the antigen-binding region, the use of carbohydrate-directed heterobifunctional crosslinking reagents, such as S-(2-thiopyridyl)-L-cysteine hydrazide (TPCH), prevents inactivation of the antigen-binding function. The synthesis of two carbohydrate-directed heterobifunctional crosslinking reagents is described. Coupling protocols for the preparation of antibody conjugates with effector proteins of different sizes using carbohydrate-directed heterobifunctional crosslinking reagents are also provided.

Antibodies↗
PubMed2002

Generation and characterization of transgenic mice expressing cobra venom factor.

Cobra venom factor (CVF), the anticomplementary protein in cobra venom, activates the alternative complement pathway, eventually leading to complement consumption. Here, we describe the development of a transgenic mouse model for CVF. We generated a DNA construct containing the full-length cDNA for single-chain pre-pro-CVF. Expression of CVF was controlled by the alpha(1)-antitrypsin promoter to achieve liver-specific expression. Linearized DNA was microinjected into murine ovary cells (strain CD(2)F(1) (BALB/cxDBA/2J)) and the newborn mice were analyzed for stable integration of CVF DNA. After establishing the transgene, mice were propagated in a BALB/c background. The CVF mRNA was detected in the liver and, in some animals, in the kidney. CVF protein was detected in small amounts in the serum. Serum complement hemolytic activity in CVF-transgenic mice was virtually absent. The concentration of plasma C3 was significantly reduced. The CVF-transgenic animals show no unusual phenotype. They provide an animal model to study the effect of long-term complement depletion by continued activation, as well as the role of complement in host immune response and pathogenesis of disease.

Animals↗