Biochemical analysis of oligomerization of expanded polyalanine repeat proteins.
Many human proteins contain amino acid repeats that can form homopolymeric amino acid (HPAA) tracts. HPAA tract proteins that contain polyalanine sequences promote diseases, including oculopharyngeal muscular dystrophy. The pathological properties of these proteins develop when the repeats match or exceed approximately 20 residues. We analyzed the oligomerization of yellow fluorescent protein (YFP) and GST fusion proteins containing >20 alanine repeats by using sucrose density gradient centrifugation. YFP and GST fusion proteins having 23 polyalanine residues sedimented readily in sucrose density gradients, suggesting instability and oligomerization of proteins with an excess of 20 alanine repeats. Moreover, GST fusion proteins were resistant to trypsin digestion after oligomerization. Oligomerized artificial proteins with long polyalanine repeats may be suitable models for studying polyalanine-related diseases.
Nojima
Jun
J
Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Tokyo, Japan.
Oma
Yoko
Y
Futai
Eugene
E
Sasagawa
Noboru
N
Kuroda
Reiko
R
Turk
Boris
B
Ishiura
Shoichi
S
D000409:Alanine / Q000235:genetics / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D019556:COS Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid / Q000235:genetics; D014162:Transfection / Q000379:methods; D014357:Trypsin / Q000494:pharmacology; D020798:Two-Hybrid System Techniques
D001426:Bacterial Proteins; D008164:Luminescent Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; C019529:polyalanine; D014357:Trypsin; D000409:Alanine