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Xylose: is a large component of plant cell walls FIGURE 13.2 Pathway Engineering for Xylose Utilization—Genes Xylose must be degraded by a specific set of reactions before its conversion to alcohol. Two genes are necessary for the initial xylose degradation, xylA and xylB. The XylA protein converts xylose to xylulose, and XylB phosphorylates this to form xylulose 5-phosphate. The two genes are carried on shuttle plasmids and transformed into bacteria such as Zymomonas. Biotechnology by Clark and Pazdernik Copyright © 2012 by Academic Press. All rights reserved. 2 FIGURE 13.6 Disruption of inaZ Prevents Ice Nucleation Cell surface proteins of P. syringae provide a nucleation point for ice. The inaZ gene encodes an icenucleating protein. Under freezing temperatures, wild-type P. syringae allow ice crystals to form, disrupting any plant tissues the bacteria are on or within. If the inaZ gene is disrupted, the P. syringae mutant will not nucleate any ice crystals, allowing the water to supercool. Biotechnology by Clark and Pazdernik Copyright © 2012 by Academic Press. All rights reserved. 3 FIGURE 13.8 Similar Reactions of Naphthalene and Indole When naphthalene (top) is attacked by an oxygenase, two hydroxyls are added to form its diol. The same oxygenase attacks the indole ring because this is similar to naphthalene. The indoxyl oxidizes spontaneously into indigo, which is blue. The indigo ring system can have alternative groups attached at the positions shown as X. If X is hydrogen, the molecule is indigo itself, but if X is bromine, the molecule becomes more purple than blue and is called Tyrian purple. Biotechnology by Clark and Pazdernik Copyright © 2012 by Academic Press. All rights reserved. 4 FIGURE 13.18 Biosynthesis of b-Lactam Antibiotics When certain molds grow on agar originally covered with bacteria, a clear zone appears around the mold where no bacteria are able to grow. The clearing is due to release of antibiotics such as penicillin and cephalosporin C from the mold. Biotechnology by Clark and Pazdernik Copyright © 2012 by Academic Press. All rights reserved. 5 FIGURE 13.19 Engineered Pathway to 7-ACA To engineer new antibacterial compounds, cephalosporin C must be converted into 7aminocephalosporanic acid (7-ACA). The enzymes involved in this conversion are d-amino acid oxidase and cephalosporin acylase. The genes for these enzymes have been isolated, cloned, and expressed in different bacteria, as well as in molds producing cephalosporin C itself. Biotechnology by Clark and Pazdernik Copyright © 2012 by Academic Press. All rights reserved. 6