N-terminal aliphatic residues dictate the structure, stability, assembly, and small molecule binding of the coiled-coil region of cartilage oligomeric matrix protein.
The coiled-coil domain of cartilage oligomeric matrix protein (COMPcc) assembles into a homopentamer that naturally recognizes the small molecule 1,25-dihydroxyvitamin D(3) (vit D). To identify the residues critical for the structure, stability, oligomerization, and binding to vit D as well as two other small molecules, all-trans-retinol (ATR) and curcumin (CCM), here we perform an alanine scanning mutagenesis study. Ten residues lining the hydrophobic pocket of COMPcc were mutated into alanine; of the mutated residues, the N-terminal aliphatic residues L37, L44, V47, and L51 are responsible for maintaining the structure and function. Furthermore, two polar residues, T40 and Q54, within the N-terminal region when converted into alanine improve the alpha-helical structure, stability, and self-assembly behavior. Helical stability, oligomerization, and binding appear to be linked in a manner in which mutations that abolish helical structure and assembly bind poorly to vit D, ATR, and CCM. These results provide not only insight into COMPcc and its functional role but also useful guidelines for the design of stable, pentameric coiled-coils capable of selectively storing and delivering various small molecules.
Gunasekar
Susheel K
SK
Department of Chemical and Biological Sciences, Polytechnic Institute of New York University, Brooklyn, New York 11201, USA.
Asnani
Mukta
M
Limbad
Chandani
C
Haghpanah
Jennifer S
JS
Hom
Wendy
W
Barra
Hanna
H
Nanda
Soumya
S
Lu
Min
M
Montclare
Jin Kim
JK
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D064236:Cartilage Oligomeric Matrix Protein; D003474:Curcumin / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007930:Leucine / Q000235:genetics; D064235:Matrilin Proteins; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D055550:Protein Stability; D017434:Protein Structure, Tertiary / Q000235:genetics; D051381:Rats; D014633:Valine / Q000235:genetics; D014801:Vitamin A / Q000235:genetics / Q000378:metabolism; D014807:Vitamin D / Q000235:genetics / Q000378:metabolism
D064236:Cartilage Oligomeric Matrix Protein; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D064235:Matrilin Proteins; D010446:Peptide Fragments; C577553:TSP5 protein, human; D014801:Vitamin A; D014807:Vitamin D; D007930:Leucine; D014633:Valine; D003474:Curcumin; D000409:Alanine