Comparative analysis of protein conformations by using 2D NOESY NMR spectra
AssigneeMomenta Pharmaceuticals, Inc.
InventorsIan Christopher Parsons, Jonathan Lansing, Carlos J. Bosques, Daniela Beccati
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Abstract
The present disclosure provides a method for determining the relative conformations of a protein provided in different protein preparations, comprising steps of: (i) obtaining a first 2D NOESY NMR spectrum of a first protein preparation; (ii) obtaining a second 2D NOESY NMR spectrum of a second protein preparation; and (iii); determining whether a protein has a different conformation in the first and second protein preparations by comparing one or more cross-peaks in the first 2D NOESY NMR spectrum with one or more corresponding cross-peaks in the second 2D NOESY NMR spectrum.
Claims34
A method of measuring differences in relative conformations of a recombinant glycoprotein provided in different preparations, comprising steps of:
- generating a first 2D NOESY NMR spectrum of a first preparation of a recombinant glycoprotein using an NMR spectrometer, wherein the recombinant glycoprotein has been recombinantly expressed in a mammalian cell;
- generating a second 2D NOESY NMR spectrum, comparable to the first spectrum, of a second preparation of the same recombinant glycoprotein using an NMR spectrometer, wherein the recombinant gycoprotein has been recombinantly expressed in a mammalian cell;
- and
- measuring differences in conformation of the recombinant glycoprotein in the first and second glycoprotein preparations by comparing one or more cross-peaks in the first 2D NOESY NMR spectrum with one or more corresponding cross-peaks in the second 2D NOESY NMR spectrum.
A method of producing a recombinant glycoprotein, comprising steps of:
- providing a first preparation of a recombinant glycoprotein from a mammalian cell culture expressing the recombinant glycoprotein;
- providing a second preparation of the same recombinant glycoprotein from a mammalian cell culture expressing the recombinant glycoprotein;
- obtaining a first 2D NOESY NMR spectrum of the first preparation;
- obtaining a second 2D NOESY NMR spectrum, comparable to the first spectrum, of the second preparation;
- determining relative conformations of the recombinant glycoprotein in the first and second glycoprotein preparations by comparing one or more cross-peaks in the first 2D NOESY NMR spectrum with one or more corresponding cross-peaks in the second 2D NOESY NMR spectrum;
- and
- producing the recombinant glycoprotein in the mammalian cell culture if the relative conformations of the recombinant glycoprotein in the first and second glycoprotein preparations are at least 75% similar.
Dependencies are inferred from the claim text.