Roche Diagnostics Operations, Inc.

Roche Diagnostics Gmbh

F. Hoffmann-la Roche Ag

Bernhard Suppmann

Harald Sobek

Rainer Mueller

Johann-Peter Thalhofer

Alfred Engel

Werner Hoelke

Michael Greif

Marco Thomann

Manfred Schmidt

Sebastian Malik

Christine Jung

Kirsten Sonn

Markus Pajatsch

Helmut Schwab

Rainer Schmuck

Bruno Frey

Bernd Schneidinger

Bernd Nidetzky

Hellmut Eckstein

Antje Liehre

Helmut Schoen

Stephanie Ronning

Ingo Curdt

Christiane Luley

Michaela Fischer

Rainer Müller

Peter Pallua

Tibor Czabany

Doris Ribitsch

Katharina Schmoelzer

Sabine Zitzenbacher

Artur Hoffmann

Thomas Marx

Hoelke Werner

The present disclosure is directed to the use of certain glycosyltransferase variants having N-terminal truncation deletions. Contrary to previous findings certain truncations were found to exhibit sialidase enzymatic activity, particularly a variant of human sialyltransferase (hST6Gal-I) with a truncation deletion involving the first 89 N-terminal amino acids of the respective wild-type polypeptide. A fundamental finding documented in the present disclosure is that there exists a variant of this enzyme which is capable of catalyzing transfer of a glycosyl moiety as well as hydrolysis thereof. Thus, disclosed is a specific exemplary variant of mammalian glycosyltransferase, nucleic acids encoding the same, methods and means for recombinantly producing the variant of mammalian glycosyltransferase and use thereof, particularly for sialylating in a quantitatively controlled manner terminal acceptor groups of glycan moieties being part of glycoproteins such as immunoglobulins.

Quantitative control of sialylation

The present disclosure is directed to the use of certain glycosyltransferase variants having N-terminal truncation deletions. It was found that the combination of two different truncation variants of human &#x3b2;-galactoside-&#x3b1;-2,6-sialyltransferase I (hST6Gal-I) exhibited different specific sialyltransferase enzymatic activities. In one example, under conditions wherein the first variant &#x394;89 hST6Gal-I catalyzed formation of bi-sialylated target molecules the second variant &#x394;108 hST6Gal-I catalyzed formation of mono-sialylated target molecules. Thus, disclosed are variants of mammalian glycosyltransferase, nucleic acids encoding the same, methods and means for recombinantly producing the variants of mammalian glycosyltransferase and use thereof, particularly for sialylating in a quantitatively controlled manner terminal acceptor groups of glycan moieties being part of glycoproteins such as immunoglobulins.

Quantitative control of sialylation and specific mono-sialylation

The present invention provides improved media for the cultivation ofand culture supernatants for the biotechnological production of collagenase enzymes. The nutrient media according to the invention comprise one or more peptones from a non-mammalian source, preferably plant-derived peptones. The media can additionally comprise fish gelatin. The invention provides media, culture supernatants comprisingcollagenase, and methods to produce said collagenase.

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The present invention provides a method for purifyingcollagenase type I and type II proteins from a complex mixture by subsequently performing a precipitation with ammonium sulfate, hydrophobic interaction chromatography, cation exchange chromatography, and anion chromatography. Conditions are provided which lead to a stabilized, partially purified preparation even after the precipitation step. The method of the invention leads to a quick and efficient removal of other proteolytic activities. The preparations according to the invention provide exceptionally pure and intact collagenase type I and type II proteins which are enzymatically active. The invention also provides blends of the two isolated proteins. The invention further provides the use of the purified collagenase proteins or blends thereof for treating a tissue sample in vitro.

Truncated terminal deoxynucleotidyl transferase (TdT) derivative from calf thymus, characterized in that the derivative in comparison to the native TdT is N-terminally truncated by up to 161 amino acids and has a 20- to 30-fold higher enzyme activity in solutions containing Coions, and its recombinant production and use.

Recombinant terminal deoxynucleotidyl transferase with improved functionality

A method is provided for the production of a bovine pancreatic protein with a specific desoxyribonuclease activity of at least 6,000 units per mg of protein. Methylotrophic yeast is used as a heterologous host organism. The bovine pancreatic protein is secreted into the growth medium from which it is purified.

Recombinant bovine pancreatic desoxyribonuclease I with high specific activity

The heterologous expression of the reverse transcriptase from the Avian Myeloblastosis Virus (AMV-RT) in prokaryotic cells and in particular() is described in the present invention. The invention also includes certain measures to simplify the purification of the heterodimeric AMV-RT.

Method for producing an active heterodimeric AMV-RT in prokaryotic cells

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Weilheim, Germany

Bernhard Suppmann