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Journal of Biochemistry Advance Access originally published online on October 19, 2007
Journal of Biochemistry 2007 142(5):627-631; doi:10.1093/jb/mvm170
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© 2007 The Japanese Biochemical Society

Immunochemical and Immunohistochemical Studies on Distribution of Elastin Fibres in Human Atherosclerotic Lesions using a Polyclonal Antibody to Elastin-derived Hexapeptide Repeat

Iori Maeda1,*, Sonoko Kishita1, Yuji Yamamoto1, Kazunari Arima2, Keiko Ideta3, Jing Meng4, Noriyuki Sakata4 and Kouji Okamoto1,{dagger}

1Department of Bioscience and Bioinformatics, Kyushu Institute of Technology, 680-4 Kawazu, Iizuka, Fukuoka 820-8502; 2Department of Chemistry and Bioscience, Faculty of Science, Kagoshima University, 1-21-35 Korimoto, Kagoshima-city, Kagoshima 890-0065; 3Institute for Material Chemistry and Engineering, Kyushu University, Kasuga, Fukuoka 816-8580; and 4Second Department of Pathology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan

*To whom correspondence should be addressed. Tel: +81-948-29-7830, Fax:+81-948-29-7801, E-mail: iori{at}bio.kyutech.ac.jb

{dagger}Correspondence may also be addressed: Tel:+81-948-29-7810, Fax:+81-948-29-7801, E-mail: okagen{at}bio.kyutech.ac.jp

Received July 7, 2007; Accepted August 29, 2007


  Abstract

A polyclonal antibody to elastin-derived hexapeptide repeat, H–(Val–Gly–Val–Ala–Pro–Gly)3–NH2, was prepared in order to investigate the differences between elastin fibres in intimal hyperplasia and media in human atheroscleroic lesions. The hexapeptide repeat and {alpha}-elastin were recognized by this polyclonal antibody in enzyme-linked immunosorbent assay (ELISA), but other elastin-derived peptides such as tetrapeptide repeat, pentapeptide repeat and nonapeptide were not. In the series of hexapeptide repeats, H–(VGVAPG)n–NH2 where n is 1–7, the polyclonal antibody reacted strongly with oligomers (n = 3–7) and weakly with dimer (n = 2), but not with monomer (n = 1). CD measurements suggested that the ß-turn structure is important for recognition by the polyclonal antibody. In an immunohistochemical study, elastin was stained more strongly in intimal hyperplasia than in media, suggesting that newly synthesized elastin in intimal hyperplasia is morphologically distinct from that in media.

Key Words: circular dichroism, elastin, immunochemistry, immunohistochemistry, polyclonal antibody

Abbreviations: BSA, bovine serum albumin; CD, circular dichroism; EDC, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide; Fmoc, 9-fluorenylmethoxycarbonyl; ELISA, enzyme-linked immunosorbent assay; FT-IR, Fourier-transform infrared spectroscopy; HBTU, 2-(1H-benzotriazole-1-yl)-1,1,3,3,-tetramethyluronium hexafluorophosphate; HOBt, 1-hydroxybenzotriazole; NMR, nuclear magnetic resonance; TFE, trifluoroethanol


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