Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF

R&D Systems | Catalog # 8329-GH

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Key Product Details

  • R&D Systems HEK293-derived Recombinant Human Lysosomal alpha-Glucosidase/GAA (8329-GH)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

HEK293

Accession Number

P10253

Applications

Enzyme Activity
View all Lysosomal alpha-Glucosidase Proteins and Enzymes »

Product Specifications

Source

Human embryonic kidney cell, HEK293-derived human Lysosomal alpha-Glucosidase protein
Ala70-Cys952, with an N-terminal 6-His tag

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

Endotoxin Level

<0.10 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

His

Predicted Molecular Mass

99 kDa

SDS-PAGE

95-105 kDa, reducing conditions

Activity

Measured by its ability to release glucose from starch.
The specific activity is >7,500 pmol/min/μg, as measured under the described conditions.

Reviewed Applications

Read 4 reviews rated 5 using 8329-GH in the following applications:

Scientific Data Images for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF

Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein Enzyme Activity Diagram.

Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein, CF (Catalog # 8329-GH) hydrolyses both alpha-1,4- and alpha-1,6-glucosidic linkages on glycogen to release terminal glucose.

Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein Enzyme Activity.

Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein, CF (Catalog # 8329-GH) is measured by its ability to release glucose from starch.

Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein SDS-PAGE.

2 μg/lane of Recombinant Human Lysosomal alpha-Glucosidase/GAA Protein (Catalog # 8329-GH) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at 95-105 kDa.

Formulation, Preparation, and Storage

8329-GH
Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
Shipping The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 6 months, -20 to -70 °C under sterile conditions after opening.

Background: Lysosomal alpha-Glucosidase

Acid alpha-glucosidase (GAA) is an enzyme that is essential in the degradation of glycogen to glucose in the lysosome (1). Defects in GAA are the cause of glycogen storage disease II, also known as Pompe's disease, which is a rare autosomal recessive metabolic disorder that damages muscle and nerve cells throughout the body, primarily due to the accumulation of glycogen in the lysosome (2). Pompe disease occurs in babies, children, and adults who inherit a defective GAA gene and affects an estimated 5,000 to 10,000 people worldwide (3). Enzyme replacement therapy (ERT) is used to treat patients with this disease (4, 5).

References

  1. Hoefsloot L.H. et al. (1988) EMBO J. 7:1697.
  2. Wan, L. et al. (2008) J. Neurol. 255:831.
  3. Fukuda, T. et al. (2007) Curr. Neurol. Neurosci. Rep. 7:71.
  4. Van Gelder, C.M. et al. (2014) J Inherit Metab Dis. In press.
  5. Toscano, A. and Schoser, B. (2013) J. Neurol. 260:951.

Long Name

Glucosidase, Alpha; Acid

Alternate Names

Acid alpha-Glucosidase, Acid Maltase, GAA, LYAG, Lysosomal alphaGlucosidase

Entrez Gene IDs

2548 (Human); 14387 (Mouse); 367562 (Rat); 102141245 (Cynomolgus Monkey)

Gene Symbol

GAA

UniProt

P10253

Additional Lysosomal alpha-Glucosidase Products

Product Documents for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF

Certificate of Analysis

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Product Specific Notices for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF

For research use only

Citations for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF

Customer Reviews for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF (4)

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  • Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF
    Name: Frank Martiniuk
    Application: Enzymatic activity in vitro
    Verified Customer | Posted 05/02/2020
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    Name: Anonymous
    Application: In vitro bioactivity in cell culture
    Verified Customer | Posted 12/01/2017
  • Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF
    Name: Anonymous
    Application: Enzymatic activity in vitro
    Verified Customer | Posted 05/23/2016
  • Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF
    Name: Anonymous
    Application: Enzymatic activity in vitro
    Verified Customer | Posted 05/17/2016

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Protocols

View specific protocols for Recombinant Human Lysosomal alpha-Glucosidase/GAA, CF (8329-GH):

Materials
  • Assay Buffer: 0.1 M Sodium Acetate, pH 4.5
  • Recombinant Human Lysosomal alpha -Glucosidase (rhGAA) (Catalog # 8329-GH)
  • Stop Solution: 4.4 mM Dinitrosalicylic Acid, 1 M Potassium Tartrate, 0.4 M Sodium Hydroxide in deionized water
  • Standard: Maltose (Sigma, Catalog # M5885), 20 mM stock in deionized water
  • Substrate: Starch from potato (Sigma, Catalog # 85642), 2% (w/v) stock in deionized water
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute 20 mM Maltose standard by adding 200 µL of the 20 mM Maltose Standard to 600 µL of Assay Buffer for a 5 mM stock. This is the first point of the standard curve.
  2. Prepare the standard curve by performing five one-half serial dilutions of the 5 mM Maltose stock in Assay Buffer. Make sure there are 400 μL in each tube for each point of the curve (remove 400 μL from the last point of the curve). Prepare one tube with only 400 μL of Assay Buffer for the curve blank. The standard curve has a range of 19.5 to 625 nmol per well.
  3. Dilute rhGAA to 32 ng/μL in Assay Buffer.
  4. Dilute 2% starch to 1% in Assay Buffer by combining equal volumes of 2% starch and Assay Buffer.
  5. Prepare reactions by combining 20 μL of diluted rhGAA with 380 μL of 1% starch (step 4). Include a control by combining 20 μL of Assay Buffer with 380 μL of 1% starch.
  6. Vortex, spin, and then incubate reactions, control and standard curve at 37 °C for 1 hour.
  7. Add 400 uL of Stop Solution to all vials, including standard curve.
  8. Heat all vials at 95-100 °C for 6 minutes. Then, cool on ice. Tip: Use lid-locks on vials when heating.
  9. Load 250 µL of each dilution of the standard curve, reactions and controls to empty wells in triplicate in plate.
  10. Read plate at 546 nm (absorbance) in endpoint mode.
  11. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

 Adjusted glucose produced* (nmol) x (1000 pmol/nmol)
Incubation time (min) x amount of enzyme (µg)

     *Derived from the maltose standard curve using linear or 4-parameter fitting and adjusted for curve blank.

Per Well:

  • rhGAA: 0.200 µg
  • Starch: 0.475%

FAQs

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