Difference between revisions of "Part:BBa K2559000"

Revision as of 14:42, 13 October 2018

Bacterial cellulose synthase Z (BcsZ) The BBa_K2559000 is an endoglucanase (Bcs Z) involved in the process of cellulose hydrolysis, and its roles in cellulose biosynthesis have long remained obscure.

Usage and Biology

Bacterial cellulose synthase Z (bcsZ) from several bacteria have been shown to possess an endo-β-1,4-glucanase (cellulase) activity, and E. coli protein was even crystallized in a complex with the substrate cellopentaose, however, the roles in cellulose biosynthesis have long remained obscure. We suspect that the BcsZ and BlgX may help the cell form the correct cellulose configuration through selective digestion.[1]

BcsZ adopts an (α/α)6-barrel fold. BcsZ is shown as a cartoon from a top (A) and side (B) view. The inner and outer helices are shown in dark and pale green, respectively, and the three β-sheets flanking the substrate binding cleft are shown in blue. The catalytic residues Glu55 and Asp243 are shown as red sticks. Helices are labeled H1–H12 from the N to the C terminus. The C-terminal residues 349–357 of BcsZ form a two-stranded β-sheet (colored orange) on the opposite side of the substrate binding pocket.[2]

Degradation of CMC by BcsZ

The BcsZ in E. coli was even crystallized in a complex with the substrate cellopentaose.When BcsZ-overexpressing E. coli cells were grown on agar plates containing 2% CMC, a clear halo appeared around the colonies after staining with Congo Red.Purified cellulase from Aspergillus niger (Tokyo Chemical Industry, GenBank entry CAA03658.1) and bovine serum albumin served as positive and negative controls, respectively.[2]

The right show degradation of CMC by BcsZ which indicate the cellulase activity of BcsZ in E.coli.

We introduced bcsZ, H, A, B, C, D, bglX from the Acetobacter xylinum which involve in the process of bacterial cellulose synthesis into the model organism cyanobacteria (Synechocystis sp.pcc6803),and we obtain the transgenic cyanobacteria.

Transgenic cyanobacteria with bcsZH-ABCD-bglX genes and the cellulose measurement

Cyanobacteria glucose content measurement (3 repeat). The same of quality of transferred cyanobacteria with bcsZH-ABCD-bglX gene and wildtype are treated with lysozyme to break out the cell. The glucose represents the content of cellulose. The addition of cellulase can digest the cellulose to glucose then the bacterial cellulose can measured. The distinct differences between the red column and blue column indicate the high expression intensity of bacteria cellulose. Wilcoxon test indicates our data is reliable.

The measurement of cellulose content in transgenic cyanobacteria which expressed bcs gene . The P-value verified that the distinction between treatment group  and control group.

The part is facilitate the in-depth research for other teams!

Reference : 1.Romling U & Galperin MY (Bacterial cellulose biosynthesis: diversity of operons, subunits, products, and functions. (Translated from eng) Trends Microbiol 23(9):545-557 (in eng). 2.Mazur O & Zimmer J (Apo- and cellopentaose-bound structures of the bacterial cellulose synthase subunit BcsZ. (Translated from eng) J Biol Chem 286(20):17601-17606 (in eng).

The proposed organization of BcsZ in the BCS complexes

Sequence and Features