structural biology, biochemistry, proteomics, microbiology, X-ray crystallography
The principal focus of my lab is to utilize proteomic and crystallographic techniques to elucidate and characterize new systems of protein complexes in Mycobacterium tuberculosis. Our overall goal is to create a systems approach to shift the focus of structural biology from a single protein to molecular assemblies. The systems of immediate interest contain potential anti-TB protein drug targets and protein membrane components. We also are studying contact-dependent growth inhibition in gram-negative bacteria.
Novel Mycobacterial Heme Uptake System
Iron is an essential metal for life. Mycobacteria must import iron from its host. Molecules involved in iron chelating pathways are well characterized, such as those involving exochelins and siderophores. Recently, it has been shown that during the early stages of S. aureus infection the major source of nutrient iron is heme rather than transferrin iron. A potential mycobacterial secreted hemophore (heme scavenging protein) has been identified in mycobacteria. Also, a potential cytosolic heme-degrading protein has been identified. Hence, a novel mcobacterial heme uptake system may exisit. My laboratory will be dissecting this pathway both structrually and biochemically. Thus far, the X-ray crystal structure of the potential hemophore has been deteremined, and a potential heme uptake membrane protein has been identified by mass spectrometry. Investigation into this uptake system is on-going.
Contact-dependent Growth Inhibition
Contact-dependent growth inhibition is a newly discovered mechanism of bacterial communication and competition. CDI toxin and immunity proteins are found in a variety of gram-negative bacteria, and are highly sequence-divergent across species. Some toxins within the CDI system require activation by forming a protein complex with a target cell protein – termed a “permissive” factor – adding a new level of complexity to the CDI system. In collaboration with the Hayes and Low labs at UCSB and MCSG, we plan
1) To solve toxin/immunity complexes that are sequence-divergent in collaboration with MCSG, in order to shed light on the requirements for toxin neutralization. Additionally, we will characterize the function of each toxin. Thus far, MCSG has cloned six toxin/immunity pairs and have two diffracting crystals, one toxin/immunity complex and one immunity protein alone. UCSB has also received these clones to biochemically characterize them.
2) To develop a screen to identify ‘permissive’ factors and biochemically characterize them. UCSB has developed a 96-well plate GFP-based assay to rapidly identify such factors.
3) To structurally characterize the protein complexes in the presence of "permissive" factors. UCI has solved the structure of a UPEC539 toxin in complex with its permissive factor CysK, which has informed functional studies.
A Single Mutation in the Mycobacterium tuberculosis Heme-Degrading Protein, MhuD, Results in Different Products.
Chao A, Goulding CW.
Biochemistry. 2019 Feb 12;58(6):489-492. doi: 10.1021/acs.biochem.8b01198.
Iron Acquisition in Mycobacterium tuberculosis.
Chao A, Sieminski PJ, Owens CP, Goulding CW.
Chem Rev. 2019 Jan 23. 119(2):1193-1220. doi: 10.1021/acs.chemrev.8b00285.
The affinity of MhuD for heme is consistent with a heme degrading function in vivo.
Thakuri B, Graves AB, Chao A, Johansen SL, Goulding CW, Liptak MD.
Metallomics. 2018 Nov 14;10(11):1560-1563
Introducing the new bacterial branch of the RNase A superfamily
Cuthbert BJ, Burley KH, Goulding CW.
RNA Biol, 2018, Jan 2;15(1):9-12. doi: 10.1080/15476286.2017.1387710.
Protein engineering: Redirecting membrane machinery.
Burley K, Goulding CW
Nat Chem Biol. 2017 Aug 18;13(9):927-928. doi: 10.1038/nchembio.2451
From B to A: making an essential cofactor in a human parasite.
Morrissette NS, Goulding CW
Biochem J. 2017 Aug 30;474(18):3089-3092. doi: 10.1042/BCJ20170446.
The CDI toxin of Yersinia kristensenii is a novel bacterial member of the RNase A superfamily
Batot G, Michalska K, Ekberg G, Irimpan EM, Joachimiak G, Jedrzejczak R, Babnigg G, Hayes CS, Joachimiak A, Goulding CW.
Nucleic Acids Res. 2017 epub Apr 10. doi: 10.1093/nar/gkx230.
Pharmacological Inhibition of Host Heme Oxygenase-1 Suppresses Mycobacterium tuberculosis Infection In Vivo by a Mechanism Dependent on T Lymphocytes.
Costa DL, Namasivayam S, Amaral EP, Arora K, Chao A, Mittereder LR, Maiga M, Boshoff HI, Barry CE 3rd, Goulding CW, Andrade BB, Sher A.
MBio, 2016, 2016 Oct 25;7(5). pii: e01675-16.
Unraveling the essential role of CysK in CDI toxin activation.
Johnson, P. M., Beck, C. M., Morse, R. P., Garza-Sánchez, F., Low, D. A., Hayes, C. S., Goulding, CW.
Proc Natl Acad. Sci, (2016) 0027-8424. doi: 10.1073/pnas.1607112113.
Functional Diversity of Cytotoxic tRNase/immunity Protein Complexes from Burkholderia pseudomallei.
Johnson, P. M., Gucinski, G. C., Garza-Sánchez, F., Wong, T., Hung, L. W., Hayes, C. S., Goulding, CW.
J Biol Chem. 2016, 0021-9258. doi: 10.1074/jbc.M116.736074
Unique coupling of mono- and dioxygenase chemistries in a single active site promotes heme degradation
Matsui T, Nambu S, Goulding CW, Takahashi S, Fukii H, Ikeda-Saito M.
Proc Natl Acad. Sci, 2016, Mar 22, epub
Salmonella Mitigates Oxidative Stress and Thrives in the Inflamed Gut by Evading Calprotectin-Mediated.
Diaz-Ochoa, V. E., Lam, D., Lee, C. S., Klaus, S., Behnsen, J., Liu, J. Z., Chim, N., Nuccio, S. P., Rathi, S. G., Mastroianni, J. R., Edwards, R. A., Jacobo, C. M., Cerasi, M., Battistoni, A., Ouellette, A. J., Goulding, CW., Chazin, W. J., Skaar, E. P., Raffatellu, M. (2016).
Cell PMID:27006503Host Microbe. 2016 Jun 8;19(6):814-25. doi: 10.1016/j.chom.2016.05.005.
Diversification of ß-augmentation interactions between CDI toxin/immunity proteins.
Morse RP, Willett JL, Johnson PM, Zheng M, Credali A, Iniguez A, Nowick JS, Hayes CS, Goulding CW
J Mol Biol. 2015, 427 (23): 3766-84
The Structure and Interactions of Periplasmic Domains of Crucial MmpL Membrane Proteins from Mycobacterium tuberculosis.
Chim N, Torres R, Liu Y, Capri J, Batot G, Whitelegge JP, Goulding CW.
Chem Biol. 2015 Aug 20;22(8):1098-107
Fe(2+) substrate transport through ferritin protein cage ion channels influences enzyme activity and biomineralization.
Behera RK, Torres R, Tosha T, Bradley JM, Goulding CW, Theil EC.
J Biol Inorg Chem. 2015 Sep;20(6):957-69
Characterization of a Mycobacterium tuberculosis nanocompartment and its potential cargo proteins.
Contreras H, Joens MS, McMath LM, Le VP, Tullius MV, Kimmey JM, Bionghi N, Horwitz MA, Fitzpatrick JA, Goulding CW.
J Biol Chem. 2014 Jun 27;289(26):18279-8
CdiA from Enterobacter cloacae delivers a toxic ribosomal RNase into target bacteria.
Beck CM, Morse RP, Cunningham DA, Iniguez A, Low DA, Goulding CW, Hayes CS.
Structure. 2014 May 6;22(5):707-18
Crystallographic and spectroscopic insights into heme degradation by Mycobacterium tuberculosis MhuD.
Graves AB, Morse RP, Chao A, Iniguez A, Goulding CW, Liptak MD.
Inorg Chem. 2014 Jun 16;53(12):5931-40
A new way to degrade heme: the Mycobacterium tuberculosis enzyme MhuD catalyzes heme degradation without generating CO.
Nambu S, Matsui T, Goulding CW, Takahashi S, Ikeda-Saito M.
J. Biol. Chem, 2013, 288(14); 10101-9.
The Mycobacterial tuberculosis Secreted Protein, Rv0203 Transfers Heme to Membrane Proteins MmpL3 and MmpL11
Owens CP, Chim N, Harmston CA, Contreras H, Iniguez A, and Goulding CW
J. Biol. Chem, 2013, 288(30); 21714-28.
The structural characterization of bacterioferritin, BfrA, from Mycobacterium tuberculosis
McMath LM, Contreras H, Owens CP and Goulding, CW
J. of Porphyrins and Phthalocyanine, 2013, 17 (3); 229-239.
Structural basis of toxicity and immunity in contact-dependent growth inhibition (CDI) systems
R. Morse, KC Nikolakakis, JLE Willett, E Gerrick, DA Low, CS Hayes & Celia W. Goulding
Proc Natl Acad. Sci, 2012, 109(25):21480-5.
Characterization of heme ligation modes of Rv0203, a secreted heme-binding protein involved in Mycobacterium tuberculosis heme uptake.
Owens CP, Du J. Dawson JD and Goulding CW.
Biochemistry, 2012, 51 (7), 1518-31
Discovery and characterization of a unique mycobacterial heme acquisition system.
Tullius MV, Harmston CA, Owens CP, Chim N, Morse RP, McMath LM, Iniguez A, Kimmey JM, Sawaya MR, Whitelegge JP, Horwitz MA, Goulding CW.
Proc Natl Acad Sci U S A. 2011, 108(12):5051-6.
Unusual Diheme Conformation of the Heme-Degrading Protein from Mycobacterium tuberculosis,
Chim N, Iniguez A, Nguyen TQ, Goulding CW,
J Mol Biol, 2010, 396(5), 595-608
Structural Biology and Molecular Biophysics
Cellular and Molecular Biosciences
Medicinal Chemistry and Pharmacology