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There is a clear need for efficient methods to produce protein therapeutics requiring mannose-termination for therapeutic efficacy. Here we report on a unique system for production of active human lysosomal acid β-glucosidase (glucocerebrosidase, GCase, EC 3.2.1.45) using seeds of the Arabidopsis thaliana complex-glycan-deficient (cgl) mutant, which are(More)
With the global spread of the pandemic H1N1 and the ongoing pandemic potential of the H5N1 subtype, the influenza virus represents one of the most alarming viruses spreading worldwide. The influenza virus sialidase is an effective drug target, and a number of inhibitors are clinically effective against the virus (zanamivir, oseltamivir, peramivir). Here we(More)
Two potent inhibitors based on the crystal structure of influenza virus sialidase have been designed. These compounds are effective inhibitors not only of the enzyme, but also of the virus in cell culture and in animal models. The results provide an example of the power of rational, computer-assisted drug design, as well as indicating significant progress(More)
The dense glycocalyx surrounding the protozoan parasite Leishmania is an essential virulence factor. It protects the parasite from hostile environments in the sandfly vector and mammalian host and supports steps of development and invasion. Therefore, new therapeutic concepts concentrate on disturbing glycocalyx biosynthesis. Deletion of genes involved in(More)
Compounds lacking oral activity may be delivered intranasally to treat influenza virus infections in mice. However, intranasal treatments greatly enhance the virulence of such virus infections. This can be partially compensated for by giving reduced virus challenge doses. These can be 100- to 1,000-fold lower than infections without such treatment and still(More)
The heterologous expression of functional mammalian integral membrane proteins still represents a significant hurdle towards their crystallization and structure elucidation. We have therefore explored the use of the OmpA signal sequence to deliberately target the expression of the murine CMP-sialic acid transporter, a Golgi-resident protein with 10 putative(More)
The enzyme mechanism of sialidase from influenza virus has been investigated by kinetic isotope methods, NMR, and a molecular dynamics simulation of the enzyme-substrate complex. Comparison of the reaction rates obtained with the synthetic substrate 4-methylumbelliferyl-N-acetyl-alpha-D-neuraminic acid and the [3,3-2H]-substituted substrate revealed(More)
Mucopolysaccharidosis (MPS) I is a lysosomal storage disease caused by a deficiency of α-L-iduronidase (IDUA) (EC 3.2.1.76); enzyme replacement therapy is the conventional treatment for this genetic disease. Arabidopsis cgl mutants are characterized by a deficiency of the activity of N-acetylglucosaminyl transferase I (EC 2.4.1.101), the first enzyme in the(More)
Mammals express the sialic acids N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) on cell surfaces, where they act as receptors for pathogens, including influenza A virus (IAV). Neu5Gc is synthesized from Neu5Ac by the enzyme cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH). In humans, this enzyme is inactive and only(More)
Rat liver Golgi and Saccharomyces cerevisiae-expressed CMP-Neu5Ac transport protein were reconstituted in phosphatidylcholine liposomes and transport of CMP-Neu5Ac into these proteoliposomes was determined. The separation of transported substrate from free substrate was performed using Multiscreen minicolumns loaded with Sephadex G-50 resin (fine). The(More)