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Recombinant Antibodies
Published in Siegfried Matzku, Rolf A. Stahel, Antibodies in Diagnosis and Therapy, 2019
Melvyn Little, Sergey M. Kipriyanov
Some of the constructs described above for making multivalent Fvs can, of course, also be used for producing multispecific Fvs. The streptavidin-scFv fusion protein, for example (Dübel et al., 1995; Kipriyanov et al., 1996a), can be made bi- or multispecific by mixing appropriate ratios of denatured scFvs together and then renaturing. Alternatively, biotinylated antibodies can be easily conjugated to an scFv-streptavidin tetramer. This procedure facilitates the production of a more homogenous bispecific product containing equal amounts of the two antibodies. Further conjugations can be made through C-terminal cysteines which can also be biotinylated for coupling to other streptavidin complexes (Kipriyanov et al., 1994). Biotinylation of the scFv can also be performed in vivo by fusing the biotin carboxyl carrier protein (BCCP) to its C-terminus (Weiss et al., 1994). Such bifunctional constructs could provide a facile means of coupling stimulatory ligands to mammalian cell surfaces. Gotter et al (1995), for example, described a simple method for anchoring ligands and proteins to the surface of cells that have been incubated for a short time with Newcastle disease virus using an scFv-streptavidin fusion protein. The scFv was directed against the haemag-glutinin-neuraminidase of the virus envelope after fusion with the cell membrane. If this single chain antibody is fused to streptavidin, biotinylated molecules can then be attached to the cell surface (unpublished results).
The Reaction Mechanism
Published in D. B. Keech, J. C. Wallace, Pyruvate Carboxylase, 2018
transcarboxylase and that the transfer is most likely to be a base on the biotin carboxyl carrier protein. Despite its rapid equilibrium with the medium, they suggest this could be the 2'0 of the ureido group of biotin.
Microbial Pathways of Lipid A Biosynthesis
Published in Helmut Brade, Steven M. Opal, Stefanie N. Vogel, David C. Morrison, Endotoxin in Health and Disease, 2020
Paul D. Rick, Christian R. H. Raetz
A functional waaM gene product is essential for cell viability above 32°C, and it appears that WaaM is somehow involved in coupling phospholipid synthesis, LPS synthesis, and growth rate. Accordingly, the LPS molecules of cells possessing an insertionally inactivated waaM gene are depleted in both laurate and myristate at both 30 and 42°C (121). However, such mutants lose viability when grown in rich media at temperatures greater than 33°C, and cell death is accompanied by filamentation and bulging at potential septa sites or at the ends of the cell (116,118). In contrast, WaaM appears to be dispensible at slow growth rates regardless of the temperature. Thus, when cells possessing an insertionally inactivated waaM gene are grown at 42°C in defined media containing a poor carbon source, generation times of greater than 70 minutes result without attendant morphological aberrations or a loss in viability (116,118). When waaM null mutants are shifted from permissive conditions (33°C, rich media) to nonpermissive conditions (42°C, rich media), they continue to grow at the same rate, but they synthesize phospholipids at the rate required for growth at 42°C, resulting in a greatly increased phospholipid:protein ratio (121). Spontaneous extragenic suppressor mutations have been isolated in accA and accB genes, which allow waaM null mutants to grow at elevated temperature in rich media (121); these genes encode the biotin carboxyl carrier protein and biotin carboxylase subunits of acetyl coenzyeme A carboxylase complex, respectively. The mutations in accA and accB result in a decreased rate of phospholipid synthesis, which apparently restores the balance between phospholipid synthesis and growth rate at elevated temperature in rich media.
Acetyl-CoA carboxylase (ACC) as a therapeutic target for metabolic syndrome and recent developments in ACC1/2 inhibitors
Published in Expert Opinion on Investigational Drugs, 2019
Leyuan Chen, Yuqing Duan, Huiqiang Wei, Hongxin Ning, Changfen Bi, Ying Zhao, Yong Qin, Yiliang Li
The structure of most eukaryotic ACCs is a polypeptide containing three major domains, biotin carboxylase (BC), biotin-containing carboxyl carrier protein (BCCP), and carboxyltransferase (CT). In most prokaryotic organisms, ACC is an enzyme complex comprised of multiple subunits. There are two isoforms of ACC found in mammals. One is ACC1 (ACCα, 265 kDa) encoded by ACACA gene on chromosome 17q12, and the other is ACC2 (ACCβ, 275 kDa) encoded by ACACB gene on chromosome 12q23 [8–11]. In mammals, ACC1 and ACC2 are highly conserved. The main difference is that ACC2 is a mitochondria protein directed by its hydrophobic N-terminal leader sequence to the mitochondrial membrane, while ACC1 is a cytoplasmic protein [12]. In liver and adipose tissue, malonyl-CoA produced by ACC1 acts as a fatty acid synthesis unit, and further extends the carbon chain to form long-chain fatty acids under the action of fatty acid synthase. ACC2 is mainly expressed in tissues such as heart and skeletal muscle, and its catalyzed production of malonyl-CoA is an inhibitor of CPT-1 (Figure 1) [13]. Due to their different distributions, selective inhibition of ACC1 or AAC2 may bring different physiological changes [11].
Mechanism of biotin carboxylase inhibition by ethyl 4-[[2-chloro-5-(phenylcarbamoyl)phenyl]sulphonylamino]benzoate
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2022
Matthew K. Craft, Grover L. Waldrop
Acetyl-CoA carboxylase (ACC) is the multi-subunit complex that catalyses the first committed step in fatty acid synthesis22. The complex catalyses the two half reactions, shown in Scheme 1. In the first half reaction, biotin carboxylase (BC) catalyses the ATP dependent carboxylation of the vitamin biotin, which is covalently attached to biotin carboxyl carrier protein (BCCP). This forms a carboxybiotin intermediate. In the second half reaction, carboxyltransferase (CT) transfers the carboxyl group from the carboxybiotin intermediate onto acetyl-CoA, forming malonyl-CoA. The BC and CT subunits retain their activity when isolated separately and can utilise free biotin as a substrate22,23.