Chemical Education Today
Reports from Other Journals
Research Advances by Angela G. King
Licorice Root Compounds May Help Fight Tooth Decay
3. Photos and a discussion of additional medicinal uses of licorice can be found in National Geographic 2006, March, p 29. 4. The American Dental Association has more information on cavity prevention available at http://www.ada.org/ (accessed Mar 2006).
Compounds isolated from licorice root may help prevent cavities, according to researchers at the University of California, Los Angeles. In test tube studies, the scientists showed that an extract from a plant root that is used to make licorice candy and other products contains at least two compounds that appear to be potent inhibitors of Streptococcus mutans, a major cause of dental caries. The two bioactive compounds are glycyrrhizol A, a new pterocarpene, and previously identified 6,8-diisoprenyl-5,7,4⬘-trihydroxyisoflavone (Figure 1). The compounds were identified in an extract of ground air-dried roots of Glycyrrhiza uralenis, or Chinese licorice. In addition to being used as flavoring and sweetening agent in candy, tobaccos, and beverages, Chinese licorice is one of the most commonly utilized traditional medicines on mainland China due to its pharmaceutical properties such as anti-ulcer, anti-inflammatory, anti-viral, and anti-carcinogenic activities. Researchers isolated the compounds using chromatographic methods, identified them through high resolution mass-spectrometry and NMR studies. The scientists then quantitatively assayed them against S. mutans. Glycyrrhizol A displayed a minimal inhibitory concentration (MIC) of 1 g/mL while that of 6,8-diisoprenyl-5,7,4⬘-trihydroxyisoflavone was 2 g/mL. More studies are needed before it is proven that the compounds effectively fight cavities in humans, caution Qing-Yi Lu (a chemist at UCLA’s School of Medicine) and Wenyuan Shi (a microbiologist at UCLA’s School of Dentistry). If further studies show promise, they say that the licorice compounds could eventually be used as cavity-fighting components in mouthwash or toothpaste.
Antibiotic Resistance In the continuing battle against antibiotic resistance, two new studies shed light on the complex defense mechanisms that pathogenic bacteria use to evade antibiotic attack, an understanding of which could lead to new, more effective antibiotics to help save lives and combat the growing problem of antibiotic resistance. The studies, both of which target chemical components in the protective membrane surrounding bacterial cells, appear in the inaugural print issue of ACS Chemical Biology, a new monthly publication of the American Chemical Society. Lipopolysaccharides: Potential Antibiotic Target Gram-negative bacteria, considered among the most virulent, include strains that are known to cause food borne illness, bubonic plague, Legionnaires’ disease, and cholera, among others. Researchers say they are among the most difficult bugs to control using antibiotics. Now a new arsenal of weapons that may be effective against Gram-negative bacteria has been identified as the result of a recent collaboration between researchers from the University of Michigan College of Pharmacy and the Borstel Research Center in Germany. Gram-negative bacteria have an outer membrane (OM) surrounding the peptidoglycan. The OM consists of two leaflets, and the outer leaflet is comprised almost exclusively of the glycolipid lipopolysaccharide (LPS, also referred to as endotoxin). LPS is the main OM surface-associated antigen and is involved in activities associated with the host immune response. The chemical structure of LPS can be divided into three regions: lipid A (embedded in the OM), an oligosaccharide core, and a hydrophilic polysaccharide chain that determines the antigenic specificity of the bacterial strain. Lipid A is the most conserved region among different species of bacteria and is the endotoxic principle of LPS. The oligosaccharide core can be further divided into inner and outer core regions, and most
More Information 1. He, Jian; Chen, Li; Heber, David; Shi, Wenyuan; Lu, QingYi. Antibacterial Compounds from Glycyrrhiza uralensis. J. Nat. Prod. 2006, 69, 121–124. 2. A discussion of licorice’s uses and history is available from Kansas State University’s Horticulture Library at http://www.oznet.ksu.edu/ library/hort2/mf2616.pdf. Additional information on cultivation of licorice can be found at http://www.oznet.ksu.edu/ksherbs/ licorice.htm (both sites accessed Mar 2006).
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HO
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HO
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Figure 1. Glycyrrhizol A (left) and 6,8diisoprenyl-5,7,4⬘-trihydroxyisoflavone (right), natural products isolated from licorice root, inhibit the bacteria associated with dental cavities. Structures provided by A. King.
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OH O
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Reports from Other Journals Gram-negative bacteria incorporate at least one 2-keto-3-deoxy-D-mannooctulosonate (Kdo) molecule into the inner core. Scientists have long believed that at least two Kdo residues must be attached to lipid A (Kdo2–lipid A) for growth, as modeled by E. coli. The biosynthesis of Kdo2–lipid A has been studied and the enzyme D-arabinose 5-phosphate isomerase (API) has been shown to initiate the biosynthetic pathway leading to Kdo2–lipid A. Previous work cloned and characterized two API genes. Now the research team, led by Ronald Woodard, has genetically engineered a strain of E. coli that lacks the ability to form Kdo. The mutant strain is viable, despite the fact that it does not
synthesize Kdo2–lipid A, and this exciting result calls into question the E. coli Kdo2-lipid A requirement. The Gram-negative bacteria LPS layer acts as a permeability barrier toward large hydrophobic molecules, and thus researchers studied the permeability of the engineered strain, KPM22 (Table 2; Figure 2). Since KPM22 utilizes lipid IVA in place of Kdo2–lipid A in its OM, scientists were interested in its vulnerability to antibiotics and detergents which are often rendered ineffective by the OM in Gram-negative bacteria. They found KPM22 supersusceptible to large hydrophobic antibiotics such as Rifampin and Erythromycin, which displayed minimum inhibitory concentra-
Table 2. Permeability Properties of KPM22
Compound
Rifampin Fusidic acid Movobiocin Erythromycin Vancomycin Bacitracinb Chloramphenicol Ampicillin Cepaloridine Sodium dodecyl sulfate (SDS) Bile saltsc
MWa (g mol⫺1)
XlogP
822.9 516.7 612.6 733.9 1449.3 1422.7 323.1 349.4 416.5
3.72 3.7 2.74 1.98 ⫺0.47 ⫺1.03 1.476 0.25 1.73
a
MIC Wildtype (mg mL⫺1)
MIC KPM22 (mg mL⫺1)
Fold Difference
16 512 256 128 256 4096 8 4 4
0.03 2 1 1 32 512 2 2 4
512 256 256 128 8 8 4 2 1
>32000 16000
8 128
>4000 125
from http://pubchem.ncbi.nlm.nih.gov/. b74000 units g⫺1. cSodium cholate and deoxycholate. Reprinted with permission from ACS Chemical Biology 2006, 1, 33–42. Copyright 2006 American Chemical Society. aData
Figure 2. Transmission electron microscopy images of wildtype (A and B) and KPM22 (C and D). IM, inner membrane; OM, outer membrane; PG, peptidoglycan; OMV, outer membrane vesicles at the surface. Scale bars = 50 nm. Reprinted with permission from ACS Chemical Biology 2006, 1, 33– 42. Copyright 2006 American Chemical Society.
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tions of 0.03 and 1 g/mL, respectively. These antibiotics have traditionally only been effective against Gram-positive bacteria, which lack an OM. “The study is further proof-of-principle that the spectrum of activity of antibiotics can be significantly extended by targeting the formation of lipopolysaccharides in the outer membrane,” says study co-author Timothy C. Meredith.
More Information 1. Meredith, Timothy C.; Aggarwal, Parag; Mamat, Uwe; Lindner, Buko; Woodard, Ronald W. Redefining the Requisite Lipopolysaccharide Structure in Escherichia coli. ACS Chem. Biol. 2006, 1, 33–42. 2. Teaching resources on microorganisms, including Gram-negative bacteria, are available at http://www.cat.cc.md.us/ courses/bio141/lecguide/ (accessed Mar 2006). 3. An experiment investigating cell membranes with Raman spectroscopy has been published. See Craig, Norman C.; Fuchsman, William H.; Lacuesta, Nanette N. Investigation of Model Cell Membranes with Raman Spectroscopy: A Biochemistry Laboratory Experiment. J. Chem. Educ. 2003, 80, 1282–1288. 4. Molecular visualizations of LPS molecules are available online at http:// www.emsl.pnl.gov/docs/mssg/www_bio/ geo.html (accessed Mar 2006).
WTA Biosynthesis Breakthrough Researchers at Harvard Medical School and Harvard University built synthetic versions of natural substrates used by TagA and TagB, key enzymes involved in the biosynthesis of wall teichoic acids (WTAs). WTAs are anionic polymeric structures in the membrane surrounding Gram-positive bacteria and are critical for survival of many organisms. WTAs are attached through a phosphodisaccharide core to peptidoglycan, the cross-linked carbohydrate polymer that is the main component of the cell wall. Many current antibiotics inhibit the biosynthesis of peptidoglycan, but since WTAs are considered essential for bacterial survival, the enzymes involved in their synthesis offer a new antibiotic target. Until recently these enzymes have been difficult to study because their substrates were
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complex, insoluble, and only available in small amounts, says Suzanne Walker, a professor in the Microbiology Department at Harvard Medical School. Before this study, the action of the enzymes could not be demonstrated in vitro due to insufficient precursors. Now Walker’s lab has combined chemical and enzymatic transformations to prepare synthetic WTA precursors from commercially available carbohydrates. The availability of synthetic precursors will make it easier to study how the Gram-positive bacterial membrane is formed and aid in the design of new antibiotics to block its formation, says Walker. Tag A and TagB catalyze the first two committed steps in WTA synthesis. The Harvard research team will soon begin screening for compounds that can block this important chemical pathway, she says. Bacillus subtilis is the model Gram-positive bacterium used in Walker’s work. Gram-positive bacteria include anthrax and other strains that cause upper respiratory infections and sepsis. In comparison to Gram-negative bacteria, they are generally considered easier to control with antibiotics. “Antibiotic resistance is a huge problem that is only going to get worse. We need new targets, especially if we’re going to circumvent resistance,” Walker says. Both research teams caution that these new approaches may be years away from human testing and clinical use. Even if the technique is effective, bacteria can eventually develop
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ways to circumvent even the best laid approaches, underscoring the need for a better understanding of resistance machinery and the availability of new antibiotics, they say. Limited and selective use of antibiotics to prevent their overuse is also a way to stem resistance, according to health experts.
More Information 1. Ginsberg, Cynthia; Zhang, Yu-Hui; Yuan, Yanqiu; Walker, Suzanne. In Vitro Reconstitution of Two Essential Steps in Wall Teichoic Acid Biosynthesis. ACS Chem. Biol. 2006, 1, 25–28. 2. This Journal has previously tied the development of antibiotics to combinatorial biosynthesis. See Pohl, Nicola L. Developing New Antibiotics with Combinatorial Biosynthesis. J. Chem. Educ. 2000, 77, 1421. 3. More information on Walker’s research can be found at http://micro.med.harvard.edu/faculty/walker.html and http:// www.chem.harvard.edu/groups/walker/index.php (both sites accessed Mar 2006). 4. Teaching resources on bacterial cell walls, including animations and self-assessment, are available at http://www.cat.cc.md.us/ courses/bio141/lecguide/ (accessed Mar 2006).
Angela G. King is Senior Lecturer in Chemistry at Wake Forest University, P.O. Box 7486, Winston-Salem, NC 27109;
[email protected].
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