Medtronic, Inc.

Restoragen, Inc.

Bionebraska, Inc.

Barton Holmquist

Daniel J Strydom

Jin Seog Seo

Fred W Wagner

Peng Luan

Yuannan Xia

Mary J Bossard

Jay S Stout

Edwin H Merrifield

Bruce E Partridge

Dennis B Henriksen

Kirk Cryer

Dan Dormady

A palladate palladium-promoted hydrolytic polypeptide cleavage process which selectively cleaves the polypeptide at a Cys-His cleavage site comprising solubilizing the polypeptide in a reaction mixture comprised of a palladate palladium promoter dissolved in a high-concentration acidic organic acid solvent.

Polypeptide cleavage process

The invention provides a process for amidating a desired peptide comprising cleaving a substrate polypeptide at a X-cysteine sequence, wherein Xis the amino acid at the peptide carboxyl-terminus and cysteine is the first amino acid of a palladium cleavage site comprising the sequence cysteine-X-X, wherein Xis any amino acid, Xis an amino acid selected from the group consisting of cysteine, histidine, or methionine, and wherein the carboxyl-terminus of the peptide is amidated upon cleavage at the X-cysteine sequence.

Composition for palladium-mediated cleavage of peptides containing CXC, CXH or CHM sequences

The invention provides methods and nucleic acid constructs to express clostripain. The source of the coding region for recombinantly expressed clostripain is

Methods and constructs for high yield expression of clostripain

A palladatepalladium-promoted hydrolytic polypeptide cleavage process which selectively cleaves the polypeptide at a Cys-His cleavage site comprising solubilizing the polypeptide in a reaction mixture comprised of a palladatepalladium promoter dissolved in a high-concentration acidic organic acid solvent.

The invention provides a palladium-promoted hydrolytic polypeptide cleavage process which selectively cleaves the polypeptide at a Cys-His cleavage site comprising solubilizing the polypeptide in a reaction mixture comprised of a palladium promoter dissolved in a high-concentration formic acid, wherein the concentration of the formic acid in the reaction mixture is between about 1 to about 22 molar.

The invention provides methods for making peptides from a polypeptide containing at least one copy of the peptide using clostripain to excise the peptide from the polypeptide. The methods enable the use of a single, highly efficient enzymatic cleavage to produce any desired peptide sequence.

Method for universal enzymatic production of bioactive peptides

The invention provides a process for amidating a desired peptide comprising cleaving a substrate polypeptide at a X-cysteine sequence, wherein X1 is the amino acid at the peptide carboxyl-terminus and cysteine is the first amino acid of a palladium cleavage site comprising the sequence cysteine-X2-X3, wherein X2 is any amino acid, X3 is an amino acid selected from the group consisting of cysteine, histidine, or methionine, and wherein the carboxylterminus of the peptide is amidated upon cleavage at the X1-cysteine sequence.

Peptide amidation process

The invention provides a method of producing a polypeptide having a C-terminal &agr;-carboxamide group. It particularly concerns an enzymatic modification of selected substrate polypeptides which result in cleavage of the substrate polypeptide to form a product peptide with a C-terminal arginine residue having an &agr;-carboxamide group (C-terminal &ldquo;Arg-NH &rdquo;). The method includes contacting an aqueous-based solution including (i) ammonia reagent and (ii) the substrate polypeptide with (iii) clostripain.

Clostripain catalyzed amidation of peptides

A method for the production of C-terminal amidated recombinant peptides is provided. The method employs a recombinant protein construct having multiple copies of a target peptide linked by intraconnecting peptides. The intraconnecting peptides permit the multicopy construct to be selectively reacted to produce product peptides having a C-terminal .alpha.-carboxamide. A recombinant gene containing a DNA sequence coding for the recombinant protein construct and an expression cassette, an expression vector and a transformed cell including the recombinant gene are also provided.

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Eagle, NE, United States of America

Barton Holmquist