University navigation
Marcelo C. Sousa Office Phone: 303 735 4341
E-mail: Marcelo.Sousa@colorado.edu
FAX: 303 492 5894
Lab: Cristol Chemistry 430
Lab Phone: 303 735 6334
Group Website:
Ph.D.: University of Buenos Aires, 1995
Pharm. D. University of Buenos Aires, 1989
Postdoctoral Fellow: School of Medicine Stanford University, 1995-2001
Structural Biology
Our laboratory utilizes biochemical and biophysical approaches, including X-Ray crystallography, to study structure and function of proteins and protein complexes. We are also interested in developing methods (cloning by recombination, ability to easily swap expression systems and affinity tags, micro-screening of expression products, etc.) to make crystallography a more efficient, high-throughput technique. Areas of current interest include the following:
The Polymixin Resistance (Pmr) Operon: Patients with Cystic Fibrosis (CF) develop chronic airway infections with the opportunistic gram negative bacteria Pseudomonas aeruginosa. These infections are characterized by airway inflammation and neutrophilic infiltration without bacterial destruction. It has been recently shown that Pseudomonas isolated from Cystic Fibrosis patients have specific, virulence-associated modifications in their lipid A structure with 4-aminoarabinose. These modifications are responsible for resistance to cationic antimicrobial peptides (CAMPs), an important component of innate immunity. The enzymes responsible for the biosynthesis of 4-aminoarabinose-lipid A are clustered in two loci termed PmrE and PmrHFIJKLM. Mutation of any of these genes except pmrM abolishes 4aminoarabinose addition to lipid A and resistance to CAMPs. We are targeting the soluble enzymes in this pathway for structural and mechanistic characterization. They are ideal targets for drug design as inhibition of the pathway for 4aminoarabinose-lipid A biosynthesis would abolish Pseudomonas resistance to antimicrobial peptides greatly enhancing the host immune response against chronic infections with Pseudomonas.
Proteins involved in the quality control mechanism of the endoplasmic reticulum (ER): Glycoproteins are synthesized in the lumen of the ER and many enter the protein export machinery to reach their final destination in the cell. The ER possesses a quality control mechanism by which aberrant, misfolded or unassembled glycoproteins are retained in the ER until they fold or are degraded. The UDP-Glc: Glycoprotein glucosyltransferase (GT) is an enzyme at the center of the above mentioned quality control mechanism as it is the folding sensor, tagging non-native proteins with a glucose residue, marking them for retention in the ER. The structural characterization of GT is a primary goal of our research program as an initial step towards understanding its mechanism.
Structural Characterization of Ca2+/Calmodulin-Dependent Protein Kinase II (CAMKII): Among the numerous Ca2+/CaM –activated enzymes, protein kinases are a prominent class of proteins that alter the function of key cellular proteins throughout the cell by phosphorylation. CaMKII, like all known Ca2+/CaM-dependent kinases, is a Ser/Thr protein kinase. Substrates phosphorylated by CaMKII are implicated in many aspects of cellular function, including the regulation of carbohydrate metabolism (glycogen synthetase and pyruvate kinase), membrane current (Ca2+, Cl-, K+ channels, and ligand-gated channels), neurotransmitter synthesis (tyrosine hydroxylase and tryptophan hydroxylase) and release (synapsin I), transcription (C/EBP- and CRE-binding protein), cytoskeletal organization (tau and microtuble-associated protein 2), intracellular calcium homeostasis (IP3 receptor, phospholamban, cardiac ryanodine receptor and Ca2+/ATPase) and long term-potentiation (AMPA receptor) and neuronal memory. CaMKII has been described as a "cognitive" kinase because of its involvement in regulating forms of learning and memory and autoregulatory properties that can be viewed as a molecular memory. Although CaMKII is ubiquitous, it is highly abundant in the brain, comprising up to 1% of total protein in the forebrain and 2% of total protein in the hippocampus, a brain region that is associated with memory. It phosphorylates proteins that modulate presynaptic synaptic transmission, as well as a host of receptors and signaling molecules in the post-synaptic cell; a process that is widely accepted as critical for synaptic plasticity. We will focus on the structure and function of CaMKII, with special emphasis on obtaining it X-Ray crystal structure.
Selected Publications
"Recognition of the Oligosaccharide and Protein Moieties of Glycoproteins by the UDP-Glc:Glycoprotein Glucosyltransferase"; Sousa, M.; Ferrero-García, M. A. and Parodi, A. J.; Biochemistry. (1992) 31; 97-105.
The Molecular Basis for the Recognition of Misfolded Glycoproteins by the UDPGlc:Glycoprotein Glucosyltransferase"; Sousa, M. C. and Parodi, A. J.; EMBO J. (1995) 14; 4196-4203
"Crystal and Solution Structures of an HslUV Protease-Chaperone Complex"; Sousa, M.C.; Trame, C. B.; Tsuruta, H.; Wilbanks, S. M.; Reddy, V. S. and McKay, D. B.; Cell. (2000) 103; 633-643.
Structure of the Universal Stress Protein of Haemophilus influenzae"; Sousa, M.C. and McKay, D. B.; Structure. (2001) 9; 1135-1141.
Crystal Structure of Skp. A Prefoldin-like Chaperone that Protects Soluble and Membrane Proteins from Aggregation"; Walton, T. A. & Sousa, M. C.; Molecular Cell. (2004) 15(3):367-74.
"Crystal Structure of E. coli ArnA (PmrI) Decarboxylase Domain. A Key Enzyme for Lipid A Modification with 4-amino-4deoxy-L-arabinose and Polymyxin Resistance"; Gatzeva-Topalova, PZ; May, A and Sousa M.C.; Biochemistry. (2004) 43(42):13370-9.
home | undergraduate | graduate | research | people | facilities | news & events | courses | alumni