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Feedstock Crop Genetic Engineering for Alcohol Fuels

Dep. of Crop and Soil Sciences, Michigan State Univ., Plant and Soil Science Bldg., East Lansing, MI 48824

View larger version (122K): [in this window] [in a new window]   Figure 1. Photo, taken in April 1933, shows a Lincoln Nebraska gas station of the Earl Coryell Co. selling "Corn Alcohol Gasoline." By permission of the Nebraska Historical Society.

View larger version (13K): [in this window] [in a new window]   Figure 2. Immunofluoresence confocal microscopy to confirm the localization of the hetrologous A. cellulolyticus endo-1,4-ß-endoglucanase E1 enzyme in transgenic maize leaf (left) compared with that of untransformed maize leaf (right). Both transgenic and the control leaves were treated with the E1 monoclonal primary antibody and the fluorescein isothiocyanate (FITC) anti-mouse secondary antibody. Green areas around cells of transgenic maize sample (left) indicate the apparent accumulation of E1 in apoplast. By permission of Current Opinion in Biotechnology (Sticklen, 2006).

View larger version (10K): [in this window] [in a new window]   Figure 3. Production of glucose from conversion of substrates including carboxymethyl cellulose (CMC), Avicel, and ammonia fiber explosion–treated corn stover (AFEX-CS) using the transgenic corn-produced heterologous A. cellulolyticus endo-1,4-ß-endoglucanase E1 enzyme. The enzymatic hydrolysis was conducted for a period of 72 h, at 50°C with 90 rpm shaking (unpublished data). These results are similar to those published in Ransom et al. (2007).

View larger version (20K): [in this window] [in a new window]   Figure 4. Comparison of percentage of feedstock crop cellulose converted into glucose through conversion of ammonia fiber explosion (AFEX)-treated and untreated (UT) corn stover (CS) and rice straw (RS) cellulose hydrolyzed using 4 mL of transgenic rice soluble proteins containing 4.9% rice E1 heterologous enzyme compared with 15 filter paper units (FPU) of Genencor commercial Spezyme CP microbial endoglucanase and exoglucanase mix (unpublished data). Novozymes commercial microbial ß-glucosidase (6.5 mg 15 mL–1) was added to both the rice E1 heterologous enzyme and to the commercial mix enzymes to inhibit cellubiose inhibition. These results are similar to those published in Oraby et al. (2007).

View larger version (19K): [in this window] [in a new window]   Figure 5. Lignin biosynthesis pathway. PAL, phenyl ammonia lyase; C4H, cinnamate 4-hydroxylase; C3H, para-coumarate 3-hydroxylase; COMT, caffeic acid O-methyltransferase; CCoAOMT, caffeoyl-CoA O-methyltransferase; 4CL, 4-coumarate:CoA ligase; 4CL??, certain species have 4CL activity toward sinapic acid; CCR, cinnamoyl-CoA reductase; CAD, cinnamyl alcohol dehydrogenase; SAD, sinapyl alcohol dehydrogenase; HCT, para-hydroxycinnamoyl-CoA:quinate shikimate para-hydroxycinnamoyltransferase; CCR? And F5H?, enzymes whose substrates have not been tested; F5H: ferulate 5-hydroxylase; ?, conversion has been demonstrated; ??, direct conversion not convincingly been demonstrated. ***, enzymatic assays in Arabidopsis have shown that the shikimate and quinate esters of para-coumaric acid are the ideal substrates for para-coumarate 3-hydroxylase (C3H). This means that in Arabidopsis, 4CL first converts the para-coumarate to para-coumaroyl-CoA, and then the C3H converts the para-coumaroyl-shikimate and para-coumaryoyl-quinate. This figure is a less-comprehensive summary of a 2003 report (Boerjan et al., 2003) and the 2002 redrawing of lignin biosynthesis pathway (Humphreys and Chapple, 2002).