Native Glutaminase from Microorganism

Products

Native Glutaminase from Microorganism

Product Specifications

Cat#	NATE-1013
Descriptions	Microbial Glutaminase efficiently hydrolyzes glutamine present in clinical serum and cell culture samples to release glutamate for downstream biochemical quantification. As a professional auxiliary diagnostic enzyme, it supports amino acid metabolism disorder detection and auxiliary clinical biochemical analysis projects with high hydrolysis efficiency and low miscellaneous enzymatic activity.
Applications	This product can be combined with related auxiliary enzymes to realize enzymatic quantification of glutamine in serum for clinical diagnosis.
Synonyms	Glutamine aminohydrolase
EC Number	EC 3.5.1.2
Catalysis	L-Glutamine + H₂O → L-Glutamic acid + NH3
Activity	≥ 800 U/g (Glutaminase activity, In-house method)
Appearance	Light brown powder
Additive	Not added
Molecular Weight	60,500 (Amino acid sequence)
Optimum pH	6.2
pH Stability	5.0-9.0 (37°C, 16 h)
Optimum Temperature	60-65°C
Thermal Stability	up to 50°C (pH 6.0, 1 h)
Unit Definition	1 unit is defined as the enzyme quantity which generates 1 μ mole of L-glutamic acid per minute under the conditions described below.
Storage	18 months from the date of analysis when stored at 20°C or below in a dry place under sealed conditions.

General Properties

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Assay Principle

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Activity Assay

Unit Definition

1 unit is defined as the enzyme quantity which generates 1 μ mole of L-glutamic acid per minute under the conditions described below.

Reagents

A. 1 M Acetic acid buffer (acetic acid-sodium acetate, pH 6.0)
B. Triton X-100 solution (50 mg/mL)
C. Sodium hydroxide solution: Weigh 3.0 g of sodium hydroxide in 80 mL of deionized water and cool. Fill up to 100 mL with deionized water.
D. Perchloric acid solution: Dilute 8.3 mL of perchloric acid (60%) to 100 mL with deionized water.
E. Diluent: Mix approx. 600 mL of deionized water, 1 mL of Triton X-100 solution (B), and 10 mL of 1 M Acetic acid buffer (A), then fill up to 1000 mL with deionized water.
F. L-Glutamic acid:
F-1: R1 enzyme reagent F-2: R2 enzyme reagent F-3: Glutamic acid standard solution (250 mg/L)
G. Substrate solution: Weigh 1.00 g of L-glutamine and dissolved in approx. 35 mL of deionized water, then add 5 mL of 1 M Acetic acid buffer (A). Fill up to 50 mL with deionized water.
H. Enzyme solution: Weigh out Glutaminase and dissolve in Diluent (E). Enzyme solution should be prepared so that the value of (AS-ASB) becomes in the range of 0.2-0.5 (0.45-1.10 U/mL).

Procedure

Glutaminase reaction: Pipette 1.0 mL of Enzyme solution (H) into a test tube and keep at 37 ± 0.5℃ for 5 minutes. Then, add 1 mL of preheated (37 ± 0.5℃) Substrate solution (G) into the test tube and mix, keep the mixture at 37 ± 0.5℃. Exactly after 10 minutes, add Perchloric acid solution (D) and mix, then place in iced water. After 1 minute or more, add 1 mL of Sodium hydroxide solution (C) and mix (Reaction solution). L-Glutamic acid Assay Kit: Pipette 1800 µL of R1 enzyme reagent (F-1) and 40 µL of Reaction solution and mix in another test tube, then pipette 1800 µL of R2 enzyme reagent (F-2) and mix. Exactly after 20 minutes at room temperature, measure the absorbances of the mixture at 555 nm (AS, Reaction mixture). As the enzyme blank, pipette 1 mL of Enzyme solution (H) in an empty test tube, add 1 mL of Perchloric acid solution (D) and mix, and keep 37 ± 0.5℃ for 5min. Add 1 mL of Substrate solution (G) and immediately place in iced water. After 1 minute or more, add 1 mL of Sodium hydroxide solution (C) and mix. After then, follow the procedure of L-Glutamic acid Assay Kit and measure the absorbance at 555nm (ASB). Also using Glutamic acid standard solution (F-3) or water, measure the absorbance following the same procedure as for the enzyme blank (AR, ARB).

Calculation

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Safe Handling Avoid inhalation; in the event of direct skin or eye contact, immediately flush the affected area with abundant water, and consult the SDS for further information.