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Mechanism of bacterial cell-surface attachment revealed by the structure of cellulosomal type II cohesin-dockerin complex

“…Crystal structures of type II Coh modules from C. thermocellum (26), Acetivibrio cellulolyticus (27), and Bacteroides cellulosolvens (28) exhibit the ␤-jellyroll topology common to the type I Coh modules. A unique crowning ␣-helix, first identified by solution NMR studies (29), and two regions disrupting strands 4 and 8, termed''␤-flaps,'' were also observed and have been proposed to play roles in the type II interaction and specificity (26,28). However, to unambiguously identify the underlying structural elements responsible for the type II interaction and type I-type II specificity, a type II Coh-Doc complex is needed.…”

Preliminary X-ray characterization and phasing of a type II cohesin domain from the cellulosome ofAcetivibrio cellulolyticus

“…Both of these helices were lacking in the known type I cohesin structures derived from the primary scaffoldins of C. thermocellum and C. cellulolyticum. In this context, it is interesting to note that only the internal helix between -strands 6 and 7 was observed in recent solution-structure studies of another type II cohesin derived from the C. thermocellum SdbA anchoring protein (Smith et al, 2002). In the present work, 13…”

“…Thus far, the crystal structures of three different type I cohesins have been elucidated from the primary cellulosomal scaffoldins of Clostridium thermocellum and C. cellulolyticum Spinelli et al, 2000;Tavares et al, 1997). In addition, the preliminary chemical shifts and secondarystructure assignment of a type II cohesin from the C. thermocellum anchoring protein SdbA have recently been reported (Smith et al, 2002). A solution structure of a C. thermocellum dockerin domain has also been published (Lytle et al, 2001).…”

Purification and crystallization of a trimodular complex comprising the type II cohesin–dockerin interaction from the cellulosome ofClostridium thermocellum

“…The refolded protein sample was applied onto a Hi-Load 16/60 Superdex 75 size-exclusion column (Amersham Pharmacia Biosciences) equilibrated with buffer B and was eluted in 2 ml fractions using the same buffer. Fractions containing protein were pooled and concentrated to a ®nal volume of 5 ml using a Millipore Amicon 10 kDa centrifugal device and were stored at 253 K. Recombinant C. thermocellum SdbA type II cohesin was expressed and puri®ed as described previously (Smith et al, 2002). Expression and puri®cation of seleno-l-methionine (SeMet) labeled SdbA type II cohesin and DocX was performed in a manner similar to that previously described for the uniform 15 N-labeling of SdbA type II cohesin (Smith et al, 2002), with the exception that in the current study the expression plasmids were transformed into the auxotrophic Escherichia coli strain DL41 and the medium was supplemented with 50 mg SeMet rather than 1 g 15 N-NH 4 Cl (Cambridge Isotope Laboratories).…”

“…While sequential comparison, mutagenesis and biophysical studies of the type I and type II cohesin and dockerin modules have provided insight into the regions responsible for the type I and type II interactions and into their speci®city (Mechaly et al, 2000(Mechaly et al, , 2001Miras et al, 2002;Handelsman et al, 2004;Jindou, Soda et al, 2004), it is recent detailed structural studies that have dramatically moved this ®eld forward. These include the crystal structures of two C. thermocellum (Shimon et al, 1997;Tavares et al, 1997) and one C. cellulotyticum type I cohesin modules (Spinelli et al, 2000), a C. thermocellum type I dockerin NMR solution structure (Lytle et al, 2001), the NMR backbone sequential assignment and secondary-structure determination of a C. thermocellum type II cohesin module (Smith et al, 2002), a crystal structure of a type II cohesin from Acetivibrio cellulolyticus (Noach et al, 2003) and the recent crystal structure of a C. thermocellum type I cohesin±dockerin complex (Carvalho et al, 2003). However, only a direct comparison of the type I complex with a type II cohesin±dockerin complex structure will unequivocally de®ne the underlying structural elements responsible for the type I/type II speci®city.…”