Galactose Oxidase from Dactylium dendroides, CAS No.9028-79-9

CAS: 9028-79-9 Cat. No.: G128460 EC Number: 232-843-7 PubChem CID: 135318662

GRADE & PURITY EnzymoPure™, ≥30 units/mg dry weight

Accession #

P0CS93

Overview

Galactose oxidase oxidizes galactose and some galactose derivatives in both free and polymeric forms. Oxidation occurs at the C6 position. The enzyme has a molecular weight of 68 ± 3 kDa, and the optimum pH is 7.0.
Useful in the determination of lactose.

Application

Galactose Oxidase from Dactylium dendroides has been used as a component for galactose oxidase treatment of arabinogalactan. It has also been used to co-immobilise with peroxidase for the preparation of a biosensor for galactose detection. Galactose oxidase may be used as an analytical tool for the specific determination of D-galactose in blood plasma, plant extracts, and phospholipids. It could be used for the characterization of terminal D-galactoside units in several polymers. It may also be useful in the determination of lactose.

1、Specificity ：GAO has a wide substrate specificity, but remarkable stereospecificity, only oxidizing D-isomers of substrates (McPherson et al. 1992). GAO will oxidize galactose and some galactose derivatives in both free and polymeric form. Oxidation occurs at the C6 position. 
2、Composition：GAO contains one Cu(II) atom yet catalyzes a two-electron transfer reaction (McPherson et al. 1992). The copper is bound by two tyrosines, and two histidines (Tyr272, Tyr495, His496, and His581). In a novel post-translational covalent modification, Tyr272 is linked by a thioether bond to cysteine (Cys228), suggesting the involvement of a tyrosine radical in the catalytic mechanism. Stabilization of the radical occurs because Tyr272 of the thioether bond is liganded to the copper, creating a stacking interaction with Trp290 (Whittaker et al. 1989, Ito et al. 1991, and Whittaker et al. 2005). The structure of the enzyme has revealed extensive beta-sheet secondary structure, consistent with the high stability of the enzyme (Kosman et al. 1974). 
Most extracellular proteins of eukaryotes are modified by glycosylation during passage through the ER and golgi, leading to greater glycosylation of extracellular than intracellular forms of a protein. Unusually, the intracellular form of GAO is more highly glycosylated (9% carbohydrate) and exhibits greater stability than the extracellular form (2% carbohydrate) (Medonca and Zancan 1988). Additionally, most proteins are modified by O- and/or N-glycosylation while GAO is only modified only by O-glycosylation (Kornfield and Kornfield 1985, and McPherson et al. 1992)
3、Molecular Characteristics：The gaoA gene contains a long open reading frame from +324 to +2507, including the mature protein-coding sequence (+521 to +2507). It also contains a long untranslated upstream region and a putative pro-sequence with a monobasic cleavage site (McPherson et al. 1992).
4、Characteristics of Galactose Oxidase：
Protein Accession Number：P0CS93；
Isoelectric point：7.75 (Theoretical)
CATH Classification：Three domains:
Class: Mainly Beta
Architecture: Sandwich, 7 Propellor
Topology: Jelly Rolls, Methylamind Dehydrogenase; Chain H, Immunoglobulin-like
Molecular Weight
68.5 kDa (calculated from translated DNA sequence and SDS-polyacrylamide gel electrophoresis, McPherson et al. 1992)
68.0 ± 3.0 kDa (determined from physical measurements, Cooper et al. 1959) 
Optimal pH：7.0 (Cooper et al. 1959)  
Extinction Coefficient：
122,480 cm-1M-1 (Theoretical)
E1%, 280 = 17.87 (Theoretical)
Inhibitors
Cyanide
Diethyldithiocarbamate
Azide
Hydroxylamine
EDTA
Applications
Quantitative determination of galactose in blood and other biological fluids (Frings and Pardue 1964, Hankin 1966, and Roth et al. 1965)
Locating galactose histochemically (Roberts and Gupta 1965)
Detecting and distinguishing glycoproteins (Itaya et al. 1975)
5、Galactose Oxidase Assay：Method
The reaction velocity is measured in a peroxidase/o-tolidine coupled system as an increase in A425 resulting from the oxidation of galactose. One unit results in a change in A425 of 1.0 per minute at 25°C and pH 6.0 under the defined conditions.
Reagents
0.1 M Potassium phosphate buffer, pH 6.0
0.5% o-tolidine. Note: o-tolidine has been reported to be carcinogenic. Handle with care.
Peroxidase. Dissolve Worthington peroxidase (Code: HPOD) at a concentration of approximately 60 u/ml in reagent grade water.
10% galactose. Allow to come to equilibrium of mutarotation by allowing to stand overnight.
Enzyme
Dissolve at a concentration of 1 mg/ml in reagent grade water. Dilute further for assay to a concentration of 0.2 - 0.5 units/ml.
Procedure
Adjust spectrophotometer to 425 nm and 25°C.
Prepare tolidine-buffer mixture by adding 0.1 ml tolidine to 12 ml 0.1 M potassium phosphate buffer pH 6.0.
Pipette into each cuvette as follows:
Tolidine-buffer solution 1.7 ml
10% Galactose 1.5 ml
Peroxidase 0.1 ml
Incubate in spectrophotometer at 25°C for 3 - 4 mintues to achieve temperature equilibration and establish blank rate, if any. Add 0.1 ml of appropriately diluted enzyme and record increase in A425/min. from initial linear portion of the curve. 

Specifications

Product Name

Galactose Oxidase from Dactylium dendroides, CAS No.9028-79-9

Synonyms

D-Galactose:oxygen 6-oxidoreductase

Grade

EnzymoPure™

Specifications & Purity

EnzymoPure™, ≥30 units/mg dry weight

Biochemical and Physiological Mechanisms

Galactose oxidase catalyzes the coversion of D-galactose to D-galacto-hexodialdose. 2-Deoxy-D-galactose, lactose, melibiose, raffinose and stachyose react with galactose oxidase in the peroxidase:o-tolidine system. Essentially no oxidation of D-glucose,

Accession #

P0CS93

CAS

9028-79-9

Enzyme Commission Number

1.1.3.9

Molecule Type

Enzyme

Storage and Shipping

Concentration

≥30 units/mg dry weight

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

Unit definition

One Unit equals a change in absorbance at 425 nm of 1.000 per minute at 25°C, pH 6.0 using a peroxidase/o-tolidine coupled assay with galactose as the substrate.

References

1. Jiajia Li, Shuang Yue, Ziyuan Gao, Wenhua Hu, Zhaoliang Liu, Guoqiang Xu, Zhen Wu, Xumin Zhang, Guolin Zhang, Fuliang Qian, Junhong Jiang, Shuang Yang.  (2023)  Novel Approach to Enriching Glycosylated RNAs: Specific Capture of GlycoRNAs via Solid-Phase Chemistry.  ANALYTICAL CHEMISTRY,      [PMID:37524653] [10.1021/acs.analchem.3c01630]

2. Ziyi Tong, Shengyan Hou, Zhenkun Zhang, Zhen Liu, Yifei Zhang.  (2024)  Biochemical approaches for decoding the information stored with metabolites.  SENSORS AND ACTUATORS B-CHEMICAL,      [PMID:] [10.1016/j.snb.2024.136618]

3. Tong Xing, Yaxin Lv, Gongqing Wu, Zhou Zhang, Wanqing Zhang, Xinping Wang, Zhuolang Chen, Weining Zhao, Felipe Conzuelo, Fangyuan Zhao.  (2025)  A novel biofuel cell based on galactose oxidase and bilirubin oxidase for efficient glycerol conversion and electricity generation.  CHEMICAL ENGINEERING JOURNAL,      [PMID:] [10.1016/j.cej.2025.163474]

4. Wanqing Zhang, Xiaodong Su, Yaxin Lv, Xuelin Zhao, Zhou Zhang, Yuntong Du, Mengyu Fan, Heshan Zhao, Matthias Rögner, Felipe Conzuelo, Weining Zhao, Mei Li, Fangyuan Zhao.  (2026)  A Biophotocathode Based on Photosystem I with Record-High Photocurrent Density Coupled with Bioelectrochemical Glycerol Oxidation.  ACS Sustainable Chemistry & Engineering,      [PMID:] [10.1021/acssuschemeng.5c13204]

5. Hongxu Zhang, Mingyuan Liu, Wenjia Tian, Ke Liu, Mengyao Hao, Hairong Yu, Weikang Sun, Leilei Guo, Xiaoxu Tan, Kaiyu Gao, Tianyi Jiang, Chuanjuan Lü, Qianjin Kang, Cuiqing Ma, Longyang Dian, Ping Xu, Chao Gao.  (2026)  Enzymatic Synthesis of 3-Hydroxypyruvate and Pyruvate from CO2-Derived C1 Compounds.  ACS Catalysis,      [PMID:] [10.1021/acscatal.5c08729]