# US 12,006,530 B2 · Streptococcal proteases

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- Number: US 12,006,530 B2
- Document ID: US12006530B2
- Status: Granted
- Kind: B2
- Application number: 16/075,059
- Filed: 2017-02-03
- Published: 2019-11-14
- Granted: 2024-06-11
- Earliest claimed: 2016-02-04
- Assignees: Genovis AB
- Applicants: Genovis AB
- Inventors: Ulrich Von Pawel-Rammingen; Christian Spoerry
- Web page: https://patentgrep.com/patent/US12006530B2

## CPC

- C12Q 1/37: involving peptidase or proteinase
- C12N 9/52: derived from bacteria {or Archaea}
- C12N 9/641: {Cysteine endopeptidases (3.4.22)}
- G01N 33/6854: {Immunoglobulins}
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

The invention relates to new Streptococcal proteases designated IgdE which display IgG degrading cysteine protease activity. The invention further relates to in vitro methods of cleaving IgG using the described Streptococcal proteases, using the methods to generate Fc and Fab fragments, and using the methods to detect IgG.

## Claims

### Claim 1

An in vitro method for the cleavage of human IgG1, the method comprising contacting human IgG1 with a purified polypeptide in vitro, wherein the purified polypeptide comprises:
(a) the amino acid sequence of SEQ ID NO: 3;
(b) a variant thereof having at least 95% identity to the amino acid sequence of SEQ ID NO: 3 and having human IgG1 cysteine protease activity; or
(c) a fragment thereof of either (a) or (b), having human IgG1 cysteine protease activity.

### Claim 2 (depends on 1)

A method according to claim 1, wherein the purified polypeptide comprises the amino acid sequence of SEQ ID NO: 3.

### Claim 3 (depends on 1)

A method according to claim 1, wherein the method comprises incubating the purified polypeptide with a sample containing human IgG1 under conditions which permit specific cysteine protease activity to occur.

### Claim 4 (depends on 1)

A method according to claim 1, which further comprises identification and/or isolation of the cleavage products.

### Claim 5 (depends on 4)

A method according to claim 4, wherein said identification and/or isolation comprises analysis by gel electrophoresis or mass spectrometry.

### Claim 6 (depends on 1)

A method according to claim 1, wherein the method is used to generate Fc and Fab fragments.

### Claim 7 (depends on 1)

A method according to claim 1, wherein the method is used to detect human IgG1.

### Claim 8 (depends on 7)

A method according to claim 7, comprising:
(i) contacting a sample with the purified polypeptide under conditions that permit the human IgG1 specific cysteine protease activity of the polypeptide; and
(ii) monitoring for the presence of human IgG1 specific cleavage fragments; wherein the presence of the specific cleavage fragments is indicative of human IgG1 in the sample.

### Claim 9 (depends on 1)

A method according to claim 1, wherein the purified polypeptide is a purified recombinant polypeptide.