λ-crystallin related to dehydroascorbate reductase in the rabbit lens

Takahiro Suzuki, Masayasu Bando, Mikako Oka, Hideo Tsukamoto, Ichiko Akatsuka, Kenji Kawai, Hajime Obazawa, Shizuko Kobayashi, Makoto Takehana

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Purpose: To evaluate the relationship of λ-crystallin to reduced nicotinamide adenine dinucleotide (NADH)-dependent dehydroascorbate (DHA) reductase found specifically in the rabbit lens. Methods: DHA reductase Fractions I-IV were separated from the λ/βL1-crystallin fraction of rabbit lens soluble protein by diethylaminoethyl (DEAE)-cellulose ion-exchange column chromatography, and then the enzyme was partially purified from Fraction II by rechromatography on the same ion-exchange column. The isolated DHA reductase fractions were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting, native isoelectric focusing and two-dimensional gel electrophoresis. Results: Using Western blot and a probe of antiserum to recombinant λ-crystallin, the main 33-kDa protein band was strongly stained in all the rabbit lens DHA reductase fractions, and most of the additional protein bands of approximately 25-30 kDa were also detectable. In the partially purified enzyme, the 33-kDa subunit alone was identified as a distinct protein band by SDS-PAGE, and a main basic protein was found at pI 7.6 by native isoelectric focusing. In addition, many bands of more acidic proteins were separated from other enzyme fractions, and protein spots corresponding to the 33 and/or approximately 25-30-kDa subunits were detected in each of the more acidic proteins by two-dimensional gel electrophoresis. Conclusion: These results suggest that λ-crystallin is closely related to the DHA reductase in the rabbit lens. The above heterogeneity of the enzyme-crystallin may arise from posttranslational modifications.

Original languageEnglish
Pages (from-to)437-443
Number of pages7
JournalJapanese Journal of Ophthalmology
Volume47
Issue number5
DOIs
Publication statusPublished - 2003 Sep

Fingerprint

glutathione dehydrogenase (ascorbate)
Crystallins
Lenses
Rabbits
Proteins
Electrophoresis, Gel, Two-Dimensional
Isoelectric Focusing
Enzymes
Sodium Dodecyl Sulfate
NAD
Polyacrylamide Gel Electrophoresis
Western Blotting
Ion Exchange
Ion Exchange Chromatography
Post Translational Protein Processing
Cellulose
Immune Sera

Keywords

  • λ-Crystallin
  • Dehydroascorbate reductase
  • Enzyme-crystallin
  • Heterogeneity
  • Rabbit lens

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Suzuki, T., Bando, M., Oka, M., Tsukamoto, H., Akatsuka, I., Kawai, K., ... Takehana, M. (2003). λ-crystallin related to dehydroascorbate reductase in the rabbit lens. Japanese Journal of Ophthalmology, 47(5), 437-443. https://doi.org/10.1016/S0021-5155(03)00140-0

λ-crystallin related to dehydroascorbate reductase in the rabbit lens. / Suzuki, Takahiro; Bando, Masayasu; Oka, Mikako; Tsukamoto, Hideo; Akatsuka, Ichiko; Kawai, Kenji; Obazawa, Hajime; Kobayashi, Shizuko; Takehana, Makoto.

In: Japanese Journal of Ophthalmology, Vol. 47, No. 5, 09.2003, p. 437-443.

Research output: Contribution to journalArticle

Suzuki, T, Bando, M, Oka, M, Tsukamoto, H, Akatsuka, I, Kawai, K, Obazawa, H, Kobayashi, S & Takehana, M 2003, 'λ-crystallin related to dehydroascorbate reductase in the rabbit lens', Japanese Journal of Ophthalmology, vol. 47, no. 5, pp. 437-443. https://doi.org/10.1016/S0021-5155(03)00140-0
Suzuki, Takahiro ; Bando, Masayasu ; Oka, Mikako ; Tsukamoto, Hideo ; Akatsuka, Ichiko ; Kawai, Kenji ; Obazawa, Hajime ; Kobayashi, Shizuko ; Takehana, Makoto. / λ-crystallin related to dehydroascorbate reductase in the rabbit lens. In: Japanese Journal of Ophthalmology. 2003 ; Vol. 47, No. 5. pp. 437-443.
@article{1939e89ca3a940639864380010c29b22,
title = "λ-crystallin related to dehydroascorbate reductase in the rabbit lens",
abstract = "Purpose: To evaluate the relationship of λ-crystallin to reduced nicotinamide adenine dinucleotide (NADH)-dependent dehydroascorbate (DHA) reductase found specifically in the rabbit lens. Methods: DHA reductase Fractions I-IV were separated from the λ/βL1-crystallin fraction of rabbit lens soluble protein by diethylaminoethyl (DEAE)-cellulose ion-exchange column chromatography, and then the enzyme was partially purified from Fraction II by rechromatography on the same ion-exchange column. The isolated DHA reductase fractions were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting, native isoelectric focusing and two-dimensional gel electrophoresis. Results: Using Western blot and a probe of antiserum to recombinant λ-crystallin, the main 33-kDa protein band was strongly stained in all the rabbit lens DHA reductase fractions, and most of the additional protein bands of approximately 25-30 kDa were also detectable. In the partially purified enzyme, the 33-kDa subunit alone was identified as a distinct protein band by SDS-PAGE, and a main basic protein was found at pI 7.6 by native isoelectric focusing. In addition, many bands of more acidic proteins were separated from other enzyme fractions, and protein spots corresponding to the 33 and/or approximately 25-30-kDa subunits were detected in each of the more acidic proteins by two-dimensional gel electrophoresis. Conclusion: These results suggest that λ-crystallin is closely related to the DHA reductase in the rabbit lens. The above heterogeneity of the enzyme-crystallin may arise from posttranslational modifications.",
keywords = "λ-Crystallin, Dehydroascorbate reductase, Enzyme-crystallin, Heterogeneity, Rabbit lens",
author = "Takahiro Suzuki and Masayasu Bando and Mikako Oka and Hideo Tsukamoto and Ichiko Akatsuka and Kenji Kawai and Hajime Obazawa and Shizuko Kobayashi and Makoto Takehana",
year = "2003",
month = "9",
doi = "10.1016/S0021-5155(03)00140-0",
language = "English",
volume = "47",
pages = "437--443",
journal = "Japanese Journal of Ophthalmology",
issn = "0021-5155",
publisher = "Springer Japan",
number = "5",

}

TY - JOUR

T1 - λ-crystallin related to dehydroascorbate reductase in the rabbit lens

AU - Suzuki, Takahiro

AU - Bando, Masayasu

AU - Oka, Mikako

AU - Tsukamoto, Hideo

AU - Akatsuka, Ichiko

AU - Kawai, Kenji

AU - Obazawa, Hajime

AU - Kobayashi, Shizuko

AU - Takehana, Makoto

PY - 2003/9

Y1 - 2003/9

N2 - Purpose: To evaluate the relationship of λ-crystallin to reduced nicotinamide adenine dinucleotide (NADH)-dependent dehydroascorbate (DHA) reductase found specifically in the rabbit lens. Methods: DHA reductase Fractions I-IV were separated from the λ/βL1-crystallin fraction of rabbit lens soluble protein by diethylaminoethyl (DEAE)-cellulose ion-exchange column chromatography, and then the enzyme was partially purified from Fraction II by rechromatography on the same ion-exchange column. The isolated DHA reductase fractions were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting, native isoelectric focusing and two-dimensional gel electrophoresis. Results: Using Western blot and a probe of antiserum to recombinant λ-crystallin, the main 33-kDa protein band was strongly stained in all the rabbit lens DHA reductase fractions, and most of the additional protein bands of approximately 25-30 kDa were also detectable. In the partially purified enzyme, the 33-kDa subunit alone was identified as a distinct protein band by SDS-PAGE, and a main basic protein was found at pI 7.6 by native isoelectric focusing. In addition, many bands of more acidic proteins were separated from other enzyme fractions, and protein spots corresponding to the 33 and/or approximately 25-30-kDa subunits were detected in each of the more acidic proteins by two-dimensional gel electrophoresis. Conclusion: These results suggest that λ-crystallin is closely related to the DHA reductase in the rabbit lens. The above heterogeneity of the enzyme-crystallin may arise from posttranslational modifications.

AB - Purpose: To evaluate the relationship of λ-crystallin to reduced nicotinamide adenine dinucleotide (NADH)-dependent dehydroascorbate (DHA) reductase found specifically in the rabbit lens. Methods: DHA reductase Fractions I-IV were separated from the λ/βL1-crystallin fraction of rabbit lens soluble protein by diethylaminoethyl (DEAE)-cellulose ion-exchange column chromatography, and then the enzyme was partially purified from Fraction II by rechromatography on the same ion-exchange column. The isolated DHA reductase fractions were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting, native isoelectric focusing and two-dimensional gel electrophoresis. Results: Using Western blot and a probe of antiserum to recombinant λ-crystallin, the main 33-kDa protein band was strongly stained in all the rabbit lens DHA reductase fractions, and most of the additional protein bands of approximately 25-30 kDa were also detectable. In the partially purified enzyme, the 33-kDa subunit alone was identified as a distinct protein band by SDS-PAGE, and a main basic protein was found at pI 7.6 by native isoelectric focusing. In addition, many bands of more acidic proteins were separated from other enzyme fractions, and protein spots corresponding to the 33 and/or approximately 25-30-kDa subunits were detected in each of the more acidic proteins by two-dimensional gel electrophoresis. Conclusion: These results suggest that λ-crystallin is closely related to the DHA reductase in the rabbit lens. The above heterogeneity of the enzyme-crystallin may arise from posttranslational modifications.

KW - λ-Crystallin

KW - Dehydroascorbate reductase

KW - Enzyme-crystallin

KW - Heterogeneity

KW - Rabbit lens

UR - http://www.scopus.com/inward/record.url?scp=0041335144&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0041335144&partnerID=8YFLogxK

U2 - 10.1016/S0021-5155(03)00140-0

DO - 10.1016/S0021-5155(03)00140-0

M3 - Article

C2 - 12967857

AN - SCOPUS:0041335144

VL - 47

SP - 437

EP - 443

JO - Japanese Journal of Ophthalmology

JF - Japanese Journal of Ophthalmology

SN - 0021-5155

IS - 5

ER -