Cellulose-binding modules from extracellular matrix proteins of Dictyostelium discoideum stalk and sheath.
Cellulose-binding modules (CBMs) of two extracellular matrix proteins, St15 and ShD, from the slime mold Dictyostelium discoideum were expressed in Escherichia coli. The expressed proteins were purified to > 98% purity by extracting inclusion bodies at pH 11.5 and refolding proteins at pH 7.5. The two refolded CBMs bound tightly to amorphous phosphoric acid swollen cellulose (PASC), but had a low affinity toward xylan. Neither protein exhibited cellulase activity. St15, the stalk-specific protein, had fourfold higher binding affinity toward microcrystalline cellulose (Avicel) than the sheath-specific ShD CBM. St15 is unusual in that it consists of a solitary CBM homologous to family IIa CBMs. Sequence analysis of ShD reveals three putative domains containing: (a) a C-terminal CBM homologous to family IIb CBMs; (b) a Pro/Thr-rich linker domain; and (c) a N-terminal Cys-rich domain. The biological functions and potential role of St15 and ShD in building extracellular matrices during D. discoideum development are discussed.
Wang
Y
Y
Department of Biological Sciences, Macquarie University, Sydney, NSW, Australia.
Slade
M B
MB
Gooley
A A
AA
Atwell
B J
BJ
Williams
K L
KL
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002480:Cellulase / Q000378:metabolism; D002482:Cellulose / Q000737:chemistry*; D003001:Cloning, Molecular; D004396:Coloring Agents / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004023:Dictyostelium / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism; D005109:Extracellular Matrix / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D012394:Rosaniline Dyes / Q000494:pharmacology; D012492:Salts / Q000494:pharmacology; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
D004396:Coloring Agents; D018076:DNA, Complementary; D011518:Proto-Oncogene Proteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012394:Rosaniline Dyes; D012492:Salts; C434672:ShD extracellular matrix protein, Dictyostelium discoideum; C434674:St15 protein, Dictyostelium discoideum; C048139:Coomassie blue; D002482:Cellulose; D002480:Cellulase