Expression and Purification of Enzymatically Active Forms of the Human Lysyl Oxidase-like Protein 4

Moon Suk Kim, Sung Su Kim, Sang Taek Jung, Jung Young Park, Han Wook Yoo, Je Sang Ko, Katalin Csiszar, Sang-Yun Choi, Youngho Kim

Research output: Contribution to journalArticle

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Abstract

The lysyl oxidase-like protein 4 (LOXL4) is the latest member of the emerging family of lysyl oxidases, several of which were shown to function as copper-dependent amine oxidases catalyzing lysine-derived cross-links in extracellular matrix proteins. LOXL4 contains four scavenger receptor cysteine-rich domains in addition to the characteristic domains of the LOX family, including the copper-binding domain, the cytokine receptor-like domain, and the residues of the lysyl-tyrosyl quinone cofactor. In an effort to assess its amine oxidase activity, we expressed LOXL4 as recombinant forms attached with hexa-histidine residues at the carboxyl terminus by using an Escherichia coli expression system. The recombinant proteins were purified with nickel-chelating affinity chromatography and converted into enzymatically active forms by stepwise dialysis. The purified LOXL4 proteins showed β-aminopropionitrile-inhibitable activity of 0.022-0.032 units/mg toward a nonpeptidyl substrate, benzylamine. These results indicate that LOXL4, with the four scavenger receptor cysteine rich domains, may also function as an active amine oxidase. Availability of the pure and active forms of LOXL4 will be significantly helpful in functional studies related to substrate specificity and crystal structure of this amine oxidase, which should provide significant insights into functional differences within the LOX family members.

Original languageEnglish
Pages (from-to)52071-52074
Number of pages4
JournalJournal of Biological Chemistry
Volume278
Issue number52
DOIs
Publication statusPublished - 2003 Dec 26

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Protein-Lysine 6-Oxidase
Purification
Amines
Oxidoreductases
Proteins
Scavenger Receptors
Cysteine
Copper
Aminopropionitrile
Amine Oxidase (Copper-Containing)
Affinity chromatography
Cytokine Receptors
Dialysis
Extracellular Matrix Proteins
Substrates
Substrate Specificity
Chelation
human LOXL4 protein
Nickel
Affinity Chromatography

ASJC Scopus subject areas

  • Biochemistry

Cite this

Expression and Purification of Enzymatically Active Forms of the Human Lysyl Oxidase-like Protein 4. / Kim, Moon Suk; Kim, Sung Su; Jung, Sang Taek; Park, Jung Young; Yoo, Han Wook; Ko, Je Sang; Csiszar, Katalin; Choi, Sang-Yun; Kim, Youngho.

In: Journal of Biological Chemistry, Vol. 278, No. 52, 26.12.2003, p. 52071-52074.

Research output: Contribution to journalArticle

Kim, Moon Suk ; Kim, Sung Su ; Jung, Sang Taek ; Park, Jung Young ; Yoo, Han Wook ; Ko, Je Sang ; Csiszar, Katalin ; Choi, Sang-Yun ; Kim, Youngho. / Expression and Purification of Enzymatically Active Forms of the Human Lysyl Oxidase-like Protein 4. In: Journal of Biological Chemistry. 2003 ; Vol. 278, No. 52. pp. 52071-52074.
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