Recombinant Human MAN2B1 Protein, CF

Catalog #: 9826-GH Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
9826-GH-050

R&D Systems Recombinant Proteins and Enzymes

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Summary Product Datasheets Carrier Free Data Images Reconstitution Calculator Background Related Research Areas

Recombinant Human MAN2B1 Protein, CF Summary

Product Specifications

Purity

>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

Endotoxin Level

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

Activity

Measured by its ability to hydrolyze 4-methylumbelliferyl-alpha -D-mannopyranoside. The specific activity is >1,500 pmol/min/μg, as measured under the described conditions.

Source

Human embryonic kidney cell, HEK293-derived human MAN2B1 protein
Gly50-Gly1011, with a C-terminal 6-His tag

Accession #

O00754-1

N-terminal Sequence
Analysis

Gly50

Predicted Molecular Mass

110 kDa

SDS-PAGE

109-127 kDa, reducing conditions

Product Datasheets

9826-GH

Product Datasheet

COA

SDS

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

9826-GH

Formulation	Supplied as a 0.2 μm filtered solution in Tris and NaCl.
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. 6 months from date of receipt, -20 to -70 °C as supplied. 3 months, -20 to -70 °C under sterile conditions after opening.

Assay Procedure

Materials

Assay Buffer: 50 mM Sodium Acetate, pH 4.5
Recombinant Human MAN2B1 (rhMAN2B1) (Catalog # 9826-GH)
Substrate: 4-Methylumbelliferyl-alpha -D-mannopyranoside (Sigma, Catalog # M3657), 50 mM stock in DMSO
F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

Dilute rhMAN2B1 to 2 ng/μL in Assay Buffer.
Dilute Substrate to 400 µM in Assay Buffer.
Load 50 μL of 2 ng/μL rhMAN2B1 into the plate, and start the reaction by adding 50 μL of 400 µM Substrate. Include a Substrate Blank containing 50 μL of Assay Buffer and 50 μL of 400 µM Substrate.
Read at excitation and emission wavelengths of 365 nm and 445 nm, respectively, (top read) in kinetic mode for 5 minutes.
Calculate specific activity:

Specific Activity (pmol/min/µg) =	Adjusted V_max* (RFU/min) x Conversion Factor** (pmol/RFU)
	amount of enzyme (µg)

*Adjusted for Substrate Blank.
**Derived using calibration standard 4-Methylumbelliferone (Sigma, Catalog # M1381).

Per Well:

rhMAN2B1: 0.1 µg
Substrate: 200 µM

Reconstitution Calculator

Background: MAN2B1

The lysosomal catabolism of N-glycans on mammalian glycoproteins occurs through the sequential exoglycosidase digestion of oligosaccharides from the non‑reducing terminus down to the carbohydrate-peptide core region (1). Included among the hydrolytic activities responsible for this oligosaccharide degradation is a broad specificity exo-alpha -mannosidase MAN2B1 that catalyzes the hydrolysis of alpha 1,2-, alpha 1,3-, and alpha 1,6-mannoside linkages present in complex, hybrid, and high mannose Asn-linked glycans (2, 3, 4). Defects in MAN2B1 result in lysosomal alpha -mannosidosis that shows symptom of mental retardation, recurrent infections, impaired hearing and Hurler-like skeletal changes (5, 6).

Long Name

Mannosidase alpha, Class 2B, Member 1

Entrez Gene IDs

4125 (Human); 17159 (Mouse); 361378 (Rat)

Alternate Names

alpha B, lysosomal; EC 3.2.1; EC 3.2.1.24; LAMAN; Lysosomal acid alpha-mannosidase; MAN2B1; MANB; Mannosidase alpha class 2B member 1; Mannosidase Alpha-B; mannosidase, alpha, class 2B, member 1

Manufacturing Specifications

Aronson, N.N. Jr. and Kuranda, M.J. (1989) FASEB J. 3:2615.
Liao, Y.F. et al. (1996) J. Biol. Chem. 271:28348.
Gotoda, Y. et al. (1989) Am. J. Hum. Genet. 63:1015.
Nebes, V.L. and Schmidt, M.C. (1994) Biochem. Biophys. Res. Commun. 200:239.
Nilssen, O, et al. (1997) Hum. Mol. Genet. 6:717.
Yu, P. et al. (2015) Genet. Med. 17:971.