SmartEnzymes GingisKHAN workflow

Above Hinge Digestion of Human IgG1

GingisKHAN (Kgp) is a cysteine protease that digests human IgG1 at a single site above the hinge, generating intact and homogenous Fab and Fc fragments.

The enzyme is used to characterize antibody-based biotherapeutics using LC-MS, to study Fc glycan analysis, bispecific antibodies, affinity and avidity effects and to identify post-translational modifications.
 

  • Reliable – digests human IgG1 above the hinge
  • Fast – generates intact Fab and Fc fragments from human IgG1 within one hour
  • Middle-level characterization of human IgG1
Human IgG1
60 min reaction
Requires mild reducing conditions. 2 mM cysteine is included.
KSCDK / THTCPPCP (above the hinge)

GingisKHAN® Lyophilized

Lyophilized enzyme for above hinge digestion of human IgG1

GingisKHAN® Fab Kit

Lyophilized enzyme and affinity resin for above hinge digestion of human IgG1 and purification of Fab fragments

About GingisKHAN®

GingisKHAN workflow

GingisKHAN is a cysteine protease that digests human IgG1 at a single site above the hinge (KSCDK / THTCPPC), generating intact Fab and Fc fragments.

GingisKHAN is a lysine-specific protease. The single digestion site is a result of the three-dimensional structure of human IgG1, making this lysine exposed for the enzyme. Additional digestion sites at exposed lysines on the Fc may appear if the N-glycans are removed. GingisKHAN requires mildly reducing conditions to be active, and optimal activity is obtained at 37°C and pH 8. The reducing agent (2 mM cysteine) is provided together with the enzyme.

GingisKHAN is purified from Porphyromonas gingivalis.

GingisKHAN® Lyophilized

Lyophilized enzyme for above hinge digestion of human IgG1.

GingisKHAN Lyophilized is available as a lyophilized powder in 2000 unit vials for digestion of up to 2 mg human IgG1. The enzyme requires mildly reducing conditions to be for active, and is provided together with 5 vials of lyophilized GingisKHAN Reducing Agent.
 

  • Flexible format for method development
  • Possible to combine enzymes for improved efficiency
  • Amenable to automated workflows
  • Ready to use – just add water!

Available Products

GingisKHAN Lyophilized 2000 unitsLyophilized enzyme for above hinge digestion of 2 mg human IgG1B0-GKH-020607.00Buy / Request a Quote

For information on how to order, visit Place an Order or contact us directly at order@genovis.com.

Unit Definition

One unit cleaves ≥ 95% of 1μg human IgG1 when incubated in 0.1 M Tris, pH 8.0 at 37°C for 1 hour.

Content and Storage

GingisKHAN Lyophilized is reconstituted by addition of water. No preservatives added. It is supplied with 5 vials of lyophilized GingisKHAN Reducing Agent (10x), containing 20 mM cysteine upon reconstitution.

GingisKHAN Lyophilized and GingisKHAN Reducing Agent are shipped cold and should be stored at -20°C upon arrival.

After reconstitution, the GingisKHAN enzyme is stable for at least 1 month at +4-8°C. Reconstituted GingisKHAN Reducing Agent cannot be stored - use it freshly prepared!

GingisKHAN® Fab Kit

Lyophilized enzyme and affinity resin for above hinge digestion of human IgG1 and purification of Fab fragments.

GingisKHAN Fab Kit contains one vial of lyophilized GingisKHAN, five vials of GingisKHAN Reducing Agent for antibody digestion, and four microspin columns of CaptureSelect™* CH1 resin for affinity purification of the Fab fragments. The CaptureSelect™ IgG-CH1 affinity matrix recognizes the CH1 domain of human IgG antibodies, enabling purification of Fab fragments independent on the light-chain isotype. Due to its unique selectivity for the CH1 domain, no co-purification of free light-chain contaminants will occur. Each CaptureSelect™ microspin column contains sufficient material to purify 0.5 mg human IgG1.
 

  • Purification of homogeneous Fab fragments from human IgG1
  • Reliable workflow for preparative applications
  • Enables chain pairing characterization of bispecific antibodies

Available Products

GingisKHAN Fab Kit 2 mgLyophilized enzyme and affinity resin for above hinge digestion of 2 mg human IgG1 and purification of fragmentsB0-GFK-0201,367.00Buy / Request a Quote

For information on how to order, visit Place an Order or contact us directly at order@genovis.com.

Unit Definition

One unit of GingisKHAN enzyme digests ≥ 95% of 1μg human IgG1 when incubated in 0.1 M Tris, pH 8.0 at 37°C for 1h.

Content and Storage

GingisKHAN Fab Kit is shipped cold. The GingisKHAN Lyophilized vials and GingisKHAN Reducing Agent should be stored at -20°C upon arrival.

The microspin columns containing CaptureSelect™* IgG-CH1 resin should be stored at +4-8°C. Do not freeze the spin columns!

After reconstitution, the GingisKHAN enzyme is stable for 2 months at +4-8°C. Reconstituted GingisKHAN Reducing Agent cannot be stored - use it freshly prepared!

Instructions

Product Specifications

Safety Data Sheet

Certificate of Analysis

* Thermo Scientific™ CaptureSelect™ resin from Thermo Fisher Scientific Inc. and its subsidiaries. Thermo Fisher and CaptureSelect are trademarks of Thermo Fisher Scientific Inc. and its subsidiaries.

FAQ and Support

Popular FAQ

Yes. GingisKHAN is provided lyophilized. After reconstitution, the enzyme can be used for up to 4 different antibodies. The digestion is performed in solution and you can prepare Fab fragments from 4 x 0.5 mg human IgG1. The purification step is done with Capture Select columns and 4 columns are provided in the kit.

Yes, GingisKHAN should be able to digest a Fc fusion protein if you have not fused the protein too close to the cleavage site and the Fc-part is from a human IgG1. GingisKHAN digests human IgG1 just above the hinge (…KSCDK / THTCPPCP…). If there are exposed Lysines in the fusion part, digestion might occur there as well.

We recommend using the 10x reducing agent provided with GingisKHAN which has an optimized pH to provide full enzymatic activity without affecting the disulphide bridges between the light chain and Fd region of the heavy chain, keeping the Fab intact. If other reducing agent is used there is a risk for non-optimal reaction conditions. It is important to use freshly prepared Reducing Agent (max storage 6h) for guaranteed activity of GinigsKHAN.

No, GingisKHAN is a general protease that with the recommended reaction conditions, 1 hour with light reducing conditions, digests human IgG1 at one specific site (…KSCDK/THTCPPCP…) above the hinge, generating Fab and Fc.

No. GingisKHAN is a protease that digests at exposed Lysines with no secondary structure restrictions. At the recommended reaction conditions, GingisKHAN digestion above the hinge has only been confirmed on human IgG1 with the exposed lysine above the hinge, (…KSCDK / THTCPPCP…). No digestion has been confirmed on other subclasses of human IgG except for several digestion sites in human IgG3.

 

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