Comparative Biochemical and Structural Analysis of Novel Cellulose Binding Proteins (Tapirins) from Extremely Thermophilic Caldicellulosiruptor Species

Vladimir Lunin, Petri Alahuhta, Yannick Bomble, Michael Himmel, Laura Lee, William Hart, Sara Blumer-Schuette, Michael Adams, Robert Kelly

Research output: Contribution to journalArticlepeer-review

12 Scopus Citations

Abstract

Genomes of extremely thermophilic Caldicellulosiruptor species encode novel cellulose binding proteins, tapirins, located proximate to the type IV pilus locus. The C-terminal domain of a tapirin (Calkro_0844) from Caldicellulosiruptor kronotskyensis is structurally unique and has a cellulose binding affinity akin to family 3 carbohydrate binding modules (CBM3). Here, full-length and C-terminal versions of tapirins from Caldicellulosiruptor bescii (Athe_1870), Caldicellulosiruptor hydrothermalis (Calhy_0908), Caldicellulosiruptor kristjanssonii (Calkr_0826), and Caldicellulosiruptor naganoensis (NA10_0869) were produced recombinantly in Escherichia coli and compared to Calkro_0844. All five tapirins bound to microcrystalline cellulose, switchgrass, poplar, filter paper, but not to xylan. Densitometry analysis of bound protein fractions visualized by SDS-PAGE revealed that Calhy_0908 and Calkr_0826 (from weakly cellulolytic species) associated with the cellulose substrates to a greater extent than Athe_1870, Calkro_0844 and NA10_0869 (from strongly cellulolytic species). Perhaps this relates to their specific needs to capture glucans released from lignocellulose by cellulases produced in Caldicellulosiruptor communities. Calkro_0844, and NA10_0869 share a high degree of amino acid sequence identity (> 80% identity), more so than with Athe_1870 (~50%). The amino acid sequence identities of Calhy_0908 and Calkr_0826 compared to Calkro_0844 were only 16% and 36%, respectively, although the three-dimensional structures of their C-terminal binding regions were closely related. Unlike the parent strain, C. bescii mutants lacking the tapirin genes did not bind to cellulose following short-term incubation, suggesting a role in cell association with plant biomass. Given the scarcity of carbohydrates in neutral terrestrial hot springs, tapirins likely help scavenge carbohydrates from lignocellulose to support growth and survival of Caldicellulosiruptor species.
Original languageAmerican English
Number of pages13
JournalApplied and Environmental Microbiology
Volume85
Issue number3
DOIs
StatePublished - 2019

NREL Publication Number

  • NREL/JA-2700-72846

Keywords

  • Caldicellulosiruptor
  • cellulase
  • glycoside hydrolase
  • lignocellulose
  • tapirins

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