Cellulolysis is severely affected in Clostridium cellulolyticum strain cipCMut1.
Progress towards understanding the molecular basis of cellulolysis by Clostridium cellulolyticm was obtained through the study of the first cellulolysis defective mutant strain, namely cipCMut1. In this mutant, a 2 659 bp insertion element, disrupts the cipC gene at the sequence encoding the seventh cohesin of the scaffoldin CipC. cipC is the first gene in a large 'cel' gene cluster, encoding several enzymatic subunits of the cellulosomes, including the processive cellulase Cel48F, which is the major component. Physiological and biochemical studies showed that the mutant strain was affected in cellulosome synthesis and severely impaired in its ability to degrade crystalline cellulose. It produced small amounts of a truncated CipC protein (P120), which had functional cohesin domains and assembled complexes which did not contain any of the enzymes encoded by genes of the 'cel' cluster. The mutant cellulolytic system was mainly composed of three proteins designated P98, P105 and P125. Their N-termini did not match any of the known cellulase sequences from C. cellulolyticum. A large amount of entire CipC produced in the cipCMut1 strain by trans-complementation with plasmid pSOScipC did not restore the cellulolytic phenotype, in spite of the assembly of a larger amount of complexes. The complexes produced in the mutant and complemented strains contained at least 12 different dockerin-containing proteins encoded by genes located outside of the 'cel' cluster. The disturbances observed in the mutant and trans-complemented strains were the result of a strong polar effect resulting from the cipC gene disruption. In conclusion, this study provided genetic evidence that the cellulases encoded by the genes located in the 'cel' cluster are essential for the building of cellulosomes efficient in crystalline cellulose degradation.
Maamar
Hédia
H
Laboratoire de Bioénergétique et Ingénierie des Protéines, UPR 9036-IBSM, CNRS, 13402 Marseille 20, France.
Valette
Odile
O
Fierobe
Henri-Pierre
HP
Bélaich
Anne
A
Bélaich
Jean-Pierre
JP
Tardif
Chantal
C
D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002480:Cellulase / Q000235:genetics* / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D003013:Clostridium / Q000235:genetics* / Q000378:metabolism*; D017931:DNA Primers; D005816:Genetic Complementation Test; D006360:Heat-Shock Proteins / Q000235:genetics; D010641:Phenotype; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction
D001426:Bacterial Proteins; C087481:ClpC protein, Bacteria; D017931:DNA Primers; D006360:Heat-Shock Proteins; D002482:Cellulose; D002480:Cellulase