Cellulose utilization by Clostridium thermocellum : Bioenergetics and hydrolysis product assimilation

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Abstract
The bioenergetics of cellulose utilization by Clostridium thermocellum was investigated. Cell yield and maintenance parameters, \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{equation*}Y_{X/{\mathrm{ATP}}}^{{\mathrm{True}}}=16.44\end{equation*} g cell/mol ATP and m = 3.27 mmol ATP/g cell per hour, were obtained from cellobiose-grown chemostats, and it was shown that one ATP is required per glucan transported. Experimentally determined values for \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{equation*}G_{{\mathrm{ATP}}}^{P-T}\end{equation*} (ATP from phosphorolytic β-glucan cleavage minus ATP for substrate transport, mol ATP/mol hexose) from chemostats fed β-glucans with degree of polymerization (DP) 2-6 agreed well with the predicted value of ( n -1)/ n ( n = mean cellodextrin DP assimilated). A mean \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{equation*}G_{{\mathrm{ATP}}}^{P-T}\end{equation*} value of 0.52 ± 0.06 was calculated for cellulose-grown chemostat cultures, corresponding to n = 4.20 ± 0.46. Determination of intracellular β-glucan radioactivity resulting from 14 C-labeled substrates showed that uptake is different for cellulose and cellobiose (G2). For 14 C-cellobiose, radioactivity was greatest for G2; substantially smaller but measurable for G1, G3, and G4; undetectable for G5 and G6; and n was ≈2. For 14 C-cellulose, radioactivity was greatest for G5; lower but substantial for G6, G2, and G1; very low for G3 and G4; and n was ≈4. These results indicate that: ( i ) C. thermocellum hydrolyzes cellulose by a different mode of action from the classical mechanism involving solubilization by cellobiohydrolase; ( ii ) bioenergetic benefits specific to growth on cellulose are realized, resulting from the efficiency of oligosaccharide uptake combined with intracellular phosphorolytic cleavage of β-glucosidic bonds; and ( iii ) these benefits exceed the bioenergetic cost of cellulase synthesis, supporting the feasibility of anaerobic biotechnological processing of cellulosic biomass without added saccharolytic enzymes.