Daniel E. GoldbergProfessor 9210 McDonnell Pediatric Research
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Research Interests | CV | Publications
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Research Interests
Parasites have evolved many clever ways to infect their hosts and develop within them. Study of these processes at a molecular level should lead to treatment or prevention of parasitic infections that afflict most of humanity. It will also shed light on general principles of biochemistry and cell biology. The organism we are studying is a protozoan parasite that causes malaria, Plasmodium falciparum.
Intraerythrocytic malaria parasites degrade vast quantities of hemoglobin to provide nutrients for their growth and maturation. This process occurs in the acidic food vacuole. My laboratory is defining the proteolytic enzymes involved, their specificities and roles in hemoglobin breakdown, as well as their targeting to the food vacuole. The data suggest an ordered catabolic pathway. Four aspartic proteases (plasmepsins), a metalloprotease (falcilysin), three cysteine proteases (falcipains), a dipeptidyl peptidase (DPAP1) and aminopeptidases are involved in the process. The biosynthesis of the aspartic proteases appears to involve targeting to the parasite surface as integral membrane proenzymes and then ingestion with their substrate hemoglobin. Once the plasmepsin precursors reach the food vacuole, they are cleaved from the membrane by the falcipains at a conserved sequence at the plasmepsin pro-mature junction.
The metalloprotease (falcilysin) cannot cleave hemoglobin or acid-denatured globin; rather it only works on fragmented globin or small, synthetic peptides. Falcilysin is capable of working at the acidic pH of the food vacuole. However, it also functions at neutral pH, where it has a very different substrate specificity. We have localized the enzyme to the apicoplast, where we believe it has a role in protein import. Further downstream in the hemoglobin degradation pathway, a dipeptidyl peptidase I of unusual specificity and multiple aminopeptidases have been isolated that appear to function in terminal degradation to liberate free amino acids.
Gene knockout studies suggest that the downstream enzymes are essential, while the upstream enzymes (plasmepsins and falcipain-2) have considerable functional overlap. Parasites can be grown in culture medium containing isoleucine as the sole exogenous amino acid, demonstrating that P falciparum can use hemoglobin as its primary amino acid source. Under these conditions, knockout growth phenotypes are apparent.
Once intraerythrocytic malaria parasites mature and replicate, they must exit the host cell to infect new erythrocytes. We have found that this is a two-step process. First, the parasites make a hole in the erythrocyte and escape bounded by their own parasitophorous vacuolar membrane. Then, the parasites must get out of this sac, which involves a specific proteolytic event. Further studies are focused on characterization of the implicated enzymes. Protease inhibitors block the escape and multiplication of the organism, suggesting that this is an attractive chemotherapeutic target.
Our work involves a combination of biochemical, genetic, genomic and physiological approaches, to gain an understanding of the biology of this nefarious organism.
keywords: Hemoglobin, Proteases, Plasmodium, Malaria, Parasitology
CV
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1979 - 1985 |
M.D. and Ph.D., Molecular Biology (Dr. Stuart Kornfeld), Washington University, St. Louis, MO |
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1978 - 1979 |
Research Assistant, Department of Biological Chemistry, laboratory of Dr. Eugene Kennedy, Harvard Medical School, Boston, MA |
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1975 - 1978 |
A.B., Harvard College, Cambridge, MA |
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Present position |
Professor of Medicine and Co-chief,
Division of Infectious Diseases |
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1997-2007 |
Director, Medical Scientist Training Program |
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1995-97 |
Associate Professor, Washington
University School of Medicine |
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1990 - 1994 |
Assistant Professor, Washington University School of Medicine |
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1988 - 1990 |
Research Associate, |
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1987 - 1988 |
Fellow in Infectious Diseases, |
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1985 - 1987 |
Resident in Internal Medicine, |
Selected Publications, Last Five Years
- Klemba M, Goldberg DE: Biological roles of proteases in parasitic protozoa. Ann Rev Biochem 2002; 71: 275-305.
- Banerjee R, Beatty W, Pelosof L, Klemba M, Goldberg DE: Four plasmepsins are active in the Plasmodium falciparum food vacuole, including a novel protease with an active site histidine. Proc Natl Acad Sci USA 2002; 99: 990-995.
- Goldberg DE: When the host is smarter than the parasite. Perspective on Zhang and Rathod, Host-parasite differences in translational autoregulation of dihydrofolate reductase, Science 2002; 296: 482-483.
- Choi C, Kim JM, Gluzman IY, Goldberg DE, Ellman JA, Marletta MA: Interference with heme binding to histidine-rich protein II as an antimalarial strategy. Chem Biol 2002; 9: 881-889.
- Akompong T, Kadekoppala M, Harrison T, Oksman A, Goldberg DE, Fujioka H, Samuel B, Sullivan D, Haldar K: Trans expression of a P. falciparum histidine-rich protein (HRP) II reveals sorting of soluble proteins in the periphery of the host erythrocyte and disrupts transport to the malarial food vacuole. J Biol Chem 2002; 277: 28923-28933.
- Ocheskey JA, Polyakov V, Oksman A, Goldberg DE, Piwnica-Worms D, Sharma V: Synthesis, characterization, and molecular structure of a gallium(III) complex of an amine-phenol ligand with activity against chloroquine-sensitive P. falciparum strains. J Inorg Biochem; 2003; 93: 265-270.
- Siripurkpong P, Yuvaniyama J, Wilairat P, Goldberg DE: Active site contribution to specificity of the aspartic proteases plasmepsins I and II. J Biol Chem 2002; 277:41009-41013.
- Harpstrite SE, Beatty AA, Collins SD, Oksman A, Goldberg DE, Sharma V: Metalloantimalarials: targeting of P. falciparum strains with novel Iron (III) and Gallium (III) complexes of an amine phenol ligand. Inorg Chem 2003; 42: 2294-2300.
- Nezami A, Kimura T, Hidaka K, Kiso A, Liu J, Kiso Y, Goldberg DE, Freire E: High affinity inhibition of a family of Plasmodium falciparum proteases by a designed adaptive inhibitor. Biochemistry 2003; 42: 8459-8464.
- Banerjee R, Francis SE, Goldberg DE: Hemoglobin-degrading plasmepsins are processed at a conserved site by an acidic convertase in Plasmodium falciparum. Mol Biochem Parasitol 2003; 129: 157-165.
- Murata C, Goldberg DE: Plasmodium falciparum falcilysin: an unprocessed food vacuole enzyme. Mol Biochem Parasitol 2003; 129: 123-126.
- Murata C, Goldberg DE: Plasmodium falciparum falcilysin: a metalloprotease with dual specificity. J Biol Chem 2003; 278: 38022-38028.
- Das TK, Samuni U, Lin Y, Goldberg DE, Rousseau DL, Friedman JM: Distal Heme Pocket Conformers of Carbonmonoxy Derivatives of Ascaris Hemoglobin: Evidence of Conformational Trapping in Porous Sol-Gel Matrices. J Biol Chem 2004; 279:10433-10441.
- Klemba M, Beatty W, Gluzman IY, Goldberg DE: Trafficking of plasmepsin II to the food vacuole of the malaria parasite Plasmodium falciparum. J Cell Biol 2004; 164: 47-56.
- Sijwali PS, Kato K, Seydel KB, Gut J, Lehman J, Klemba M, Goldberg DE, Miller LH, Rosenthal PJ: The Plasmodium falciparum cysteine protease falcipain-1 is not essential in erythrocyte stage malaria parasites. Proc Natl Acad Sci USA 2004; 101: 8721-8726.
- Klemba M, Gluzman IY, Goldberg DE: A Plasmodium falciparum dipeptidyl aminopeptidase participates in vacuolar hemoglobin degradation. J Biol Chem 2004; 279: 43000-43007.
- Liu J, Drew M, Bukanova E, Gluzman IY, Goldberg DE: The role of Plasmodium falciparum food vacuole plasmepsins. J Biol Chem 2005; 280: 1432-1437.
- Ocheskey JA, Harpstrite S, Oksman A, Goldberg DE, Sharma V: Metalloantimalarials: synthesis and characterization of a novel agent possessing activity against Plasmodium falciparum. Chem Comm 2005; 21: 1622-1624.
- Parikh S, Gut J, Istvan ES, Goldberg DE, Havlir DV and Rosenthal PJ: Antimalarial activity of HIV-1 protease inhibitors. Antimicrob Agents Chemother. 2005; 49: 2983-2985.
- Istvan EI, Goldberg DE: Distal substrate interactions enhance aspartic protease action. J Biol Chem 2005; 280: 6890-6896.
- Klemba M, Gluzman IY, Goldberg DE: Characterization of plasmepsin V, a membrane-bound aspartic protease in the endoplasmic reticulum of Plasmodium falciparum. Mol Biochem Parasitol 2005; 143: 183-191.
- Goldberg DE: Hemoglobin degradation. In Current Topics in Microbiology and Immunology, Vol. on Antimalarial Chemotherapy, Sullivan DJ, Krishna S, eds, Springer-Verlag, Heidelberg 2005; 275-291.
- Shapiro T, Goldberg DE: Drugs used in the chemotherapy of protozoal infections: Malaria. In Goodman and Gilmans The Pharmacological Basis of Therapeutics, 11th ed, Brunton LL, Parker K, Lazo J (eds), McGraw-Hill 2005; ch 39.
- Hof F, Schutz A, Fah C, Mayer S, Bur D, Liu J, Goldberg DE, Diederich F: Starving the malaria parasite: a new class of inhibitors active against the aspartic proteases plasmepsins I, II, and IV; Angew Chem 2006, 45: 2138-2141.
- Parikh S, Liu J, Sijwali P, Gut J, Goldberg DE, Rosenthal PJ: The antimalarial effects of HIV-1 protease inhibitors differ from those of the aspartic protease inhibitor pepstatin; Antimicrob Agents Chemother, 2006, 50: 2207-2209.
- Liu J, Istvan ES, Gluzman IY, Gross J, Goldberg DE: Plasmodium falciparum ensures its amino acid supply with multiple acquisition pathways and redundant proteolytic enzyme systems; Proc Natl Acad Sci USA 2006, 103: 8840-8845.
- Liu J, Istvan ES, Goldberg DE: Hemoglobin-degrading plasmepsin II is active as a monomer; J Biol Chem 2006, 281: 38682-38688.
- Ponpuak M, Klemba M, Gluzman IY, Goldberg DE: A role for falcilysin in transit peptide degradation in the Plasmodium falciparum apicoplast; Mol Micro 2007, 62: 314-334.
- Klonis N, Tan O, Jackson K, Goldberg D, Klemba M, Tilley L: Evaluation of pH during cytostomal endocytosis and vacuolar catabolism of hemoglobin by Plasmodium falciparum; Biochem J, in press.
- Armstrong CA, Goldberg DE: An FKBP destabilization domain modulates protein levels in Plasmodium falciparum; Nature Methods, in press.
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Lab Members |
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Research Assistant Professor |
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Eva Istvan |
PhD University of Texas Southwestern |
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Post-doctoral Fellows |
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Mark Drew |
PhD Johns Hopkins University |
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Ilaria Russo |
PhD University of Padua |
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Chris Armstrong |
PhD MIT |
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Vasant Muralidharan |
PhD Rockefeller University |
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Kevin Ge |
PhD University of Hong Kong |
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Aaron Miller |
MD Yale University |
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Students |
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Ericka Ricaldez |
PhD |
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Shalon Babbitt |
PhD |
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Tamira Butler |
PhD |
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Sha Sha Lu |
BA |
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Technicians |
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Anna Oksman |
MS |
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Ilya Gluzman |
PhD/DVM |
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Barb Vaupel |
MS |
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