New England Biolabs, Inc.

Ellen P Guthrie

Christopher H Taron

Chudi Guan

Phillips W Robbins

David Landry

Sharon Wong-Madden

Theodore B Davis

Ming-Qun Xu

Jack S Benner, Ii

Marta M Meda

Elizabeth M Van Cott

John Buswell

Sharon T Wong-Madden

Paula Magnelli

Dimitris Koutsioulis

Compositions and methods are provided for efficiently preparing a completely deglycosylated antibody where efficiency is measured in relative amounts of reagents in soluble or lyophilized form, and time and temperature of the reaction. Compositions and methods are also provided for separating substantially all N-linked glycans from a glycosylated antibody and for preserving functionality of the antibody. The methods are compatible with glycan labeling and protease digestion without the need for prior purification steps.

Deglycosylation reagents and methods

Methods and compositions have been described that relate to a newly identified polypeptide family wherein each member has O-glycosidase activity and specified sequence characteristics. This family of enzymes can be used for example for cleaving O-linked glycans and for synthesis of neoglycopeptides or neoglycoproteins.

[object Object]

Compositions for an alkaline phosphatase and methods for over-expression and purification of thermolabile Antarctic phosphatase (TAP) are provided. Uses for TAP include dephosphorylation of nucleic acids, sugars, peptides and proteins. TAP as described herein has advantages over phosphatases from other sources with respect to thermolability at 65&#xb0; C. and efficiency of dephosphorylation activity at approximately neutral pH.

Overexpression, purification and characterization of a thermolabile phosphatase

Substantially pure glycosidases capable for cleaving selected glycosidic bonds have been described including glycosidases isolated fromand recombinant glycosidases. Substrate specificity of isolated enzymes have been identified for GlcNac&#x3b2;1-x, Gal&#x3b1;1-3R, Gal&#x3b1;1-6R, Gal&#x3b2;1-3R, Fuc&#x3b1;-2R, Fuc&#x3b1;1-3R, Fuc&#x3b1;1-4R, Man&#x3b1;1-2R, Man&#x3b1;1-3R, Man&#x3b1;1-6R, Man&#x3b2;1-4R, Xyl&#x3b2;1-2R, Glc&#x3b2;1-4R, and Gal&#x3b2;1-4R providing improved capability for selectively cleaving a glycosidic linkage in a carbohydrate substrate and for forming modified carbohydrates.

Isolation and composition of novel glycosidases

Substantially pure glycosidases capable for cleaving selected glycosidic bonds have been described including glycosidases isolated from Xanthomonas and recombinant glycosidases. Substrate specificity of isolated enzymes have been identified for GlcNac&bgr;1-X, Gal&agr;1-3R, Gal&agr;1-6R, Gal&bgr;1-3R, Fuc&agr;-2R, Fuc&agr;1-3R, Fuc&agr;1-4R, Man&agr;1-2R, Man&agr;1-3R, Man&agr;1-6R, Man&bgr;1-4R, Xyl&bgr;1-2R, Glc&bgr;1-4R, and Gal&bgr;1-4R providing improved capability for selectively cleaving a glycosidic linkage in a carbohydrate substrate and for forming modified carbohydrates.

Substantially pure glycosidases capable for cleaving selected glycosidic bonds have been described including glycosidases isolated from Xanthomonas and recombinant glycosidases. Substrate specificity of isolated enzymes have been identified for GlcNac&bgr;1-X, Gal&agr;1-3R, Gal&agr;1-6R, Gal&bgr;1-3R, Fuc&agr;1-2R, Fuc&agr;1-3R, Fuc&agr;1-4R, Man&agr;1-2R, Man&agr;1-3R, Man&agr;1-6R, Man&bgr;1-4R, Xyl&bgr;1-2R, Glc&bgr;1-4R, and Gal&bgr;1-4R providing improved capability for selectively cleaving a glycosidic linkage in a carbohydrate substrate and for forming modified carbohydrates.

The present invention is directed to a method for cloning and producing the NaeI restriction endonuclease by 1) introducing the restriction endonuclease gene from Nocardia aerocolonigenes into a host whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid encoding and expressing the NaeI restriction endonuclease activity, and 3) purifying the NaeI restriction endonuclease from the fermented host which contains the plasmid encoding and expressing the NaeI restriction endonuclease activity.

Method for cloning and producing the NaeI restriction endonuclease and

The present invention is directed to a method for cloning and producing the SacII restriction endonuclease by 1) introducing the restriction endonuclease gene from Streptomyces achromogenes into a host whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid encoding and expressing the SacII restriction endonuclease activity, and 3) purifying the SacII restriction endonuclease from the fermented host which contains the plasmid encoding and expressing the SacII restriction endonuclease activity.

Method for producing and cloning SacII restriction endonuclease and

The present invention is directed to a method for cloning and producing the SphI restriction endonuclease by 1) introducing the restriction endonuclease gene from Streptomyces phaeochromogenes into a host whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid encoding and expressing the SphI restriction endonuclease activity, and 3) purifying the SphI restriction endonuclease from the fermented host which contains the plasmid encoding and expressing the SphI restriction endonuclease activity.

Growing community of inventors

Andover, MA, United States of America

Ellen P Guthrie