Alam, Steve
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Research Interests

My group's research interests are focused on the structural basis for protein and nucleic acid function. I have scientific interests in three areas: 1) structural investigation of proteins, peptides, and RNAs involved in protein translation or recoding (alternative-decoding); 2) biophysical studies of protein factors that sense cellular signals and organize the assembly of large protein machinery; 3) and investigation of dynamic contributions to protein function. A broad range of techniques will be used in each study including structural studies that depend upon nuclear magnetic resonance (NMR).

How does a new class of zinc-binding proteins recruit protein complex assembly? Zinc containing proteins (zinc-fingers) are one of the most abundant classes of proteins in eukaryotic genomes. Functions within this class of proteins are extraordinarily diverse; including: DNA recognition, RNA packaging, transcriptional activation, regulation of apoptosis, protein folding, assembly of protein machinery, and lipid binding. The current zinc-domains investigated contain from one to nine zinc ions and adopt five known protein folds adapting various combinations of protein side chains for Zn-coordination. Metal coordination has little functional significance and acts primarily as a folding nucleus to stabilize the folded domains. Despite this, folding of these domains can be regulated to some extent by the oxidative state of the cell through cysteine coordinating residues. In proteins containing multiple zinc-fingers, each finger can functionally become additive (as is the case with the DNA binding properties of many transcription factors), or may have adapted divergent functions (as observed in the DNA binding and protein-binding properties of the nine zinc fingers in the FOG protein (friend of GATA)). It is surprising that a large number of putative zinc-binding motifs have been identified but only a few have been structurally characterized. My group would like to contribute to the structural and functional knowledge of a new zinc-motif (UBZ; Ubiquitin Binding Zinc-motif) involved in protein machinery recruitment required by diverse cellular pathways.How does the ribosome-recycling factor disassemble a terminated ribosome? How a ribosome terminates protein synthesis and recycles translation components to continue with protein synthesis is a very basic, but poorly understood process. Termination of protein synthesis and the release of polypeptide products are signaled by a stop-codon in the ribosomal A-site. In Eubacteria, release factors (RF1, RF2 and RF3) recognize the stop-codon(s), trigger peptidyl-tRNA hydrolysis and peptide release, and dissociate from the ribosome in a GTP dependent manner requiring EF-G. Following termination, the ribosome is left in a “post-termination” state composed of the 70S ribosome, the RNA message, the final P-site deacylated-tRNA and an empty A-site. Ribosomal release factor (RRF) orchestrates the disassembly of the post-termination complex, again requiring EF-G. 70S (or 50S) dissociation by RRF promotes a “recycling” of the 30S and 50S subunits and allows translation to initiate on other RNA-messages; both of which are required for cell growth and efficient protein synthesis. Structures of ribosome recycling factors indicate RRF proteins adopt an L-shape consisting of a three-helix coil and a b/a/b sandwich domain separated by two short linker loops. A similar shape to tRNA suggests RRF functions through tRNA mimicry. Ribosome recycling proteins are found in prokaryotes and their vestiges (chloroplasts and mitochondria), making ribosome recycling an ideal anti-microbial target. Although structures of RRF are known, little is known about the structural features that are responsible for the recycling action. It has been suggested by genetic studies that ribosome recycling is dependent upon the structural arrangement or dynamic properties of the linker (or “hinge” regions) between the two domains. My groups work will be aimed at describing the physical attributes of RRF and its interactions with EF-G that govern ribosome recycling.

Publications

Steven L. Alam, Ji Sun, Marielle Payne, Brett D. Welch, B. Kelly Blake, Darrell R. Davis, Hemmo H. Meyer, Scott. D. Emr, Wesley I. Sundquist. “Ubiquitin Interactions of NZF Zinc Fingers”. Submitted Science 9/2003.

RD Fisher, B Wang, SL Alam, DS Higginson, H Robinson, WI Sundquist, CP Hill, “Structure and ubiquitin binding of the ubiquitin-interacting motif” J Biol Chem. 2003 Aug 1;278(31):28976-84.

Steven L. Alam*, Bin Wang*, Timothy L. Stemmler, Rebecca L. Rich, David G. Myszka, Wesley I. Sundquist, Hemmo H. Meyer, and Graham Warren “Structure and Ubiquitin Interaction of the NPL4 UBZ domain” J Biol Chem. 2003 May 30;278(22):20225-34. Epub 2003 Mar 18.

B. Kelly Blake, Koichi Ito, Yoshikazu Nakamura and Steven L. Alam “Letter to the editor: Backbone 1H, 13C, and 15N Assignments of the ribosome recycling factor from Thermus thermophilus” (2002) JBNMR, 24(1) 81-82.

Steven L. Alam*, Owen W. Pornillos*, Darrell R. Davis, Wesley I. Sundquist “Structure of the Tsg101 UEV domain in complex with a HIV-1 PTAP ‘late domain’ peptide”. Online Oct 15th Nature Structural Biology 2002.

T.L. Stemmler, S. L. ALAM, H. Huang, D.R. Davis, and W.I. Sundquist “A b-hairpin Switch in the Proteolytic Maturation of HIV-1 Gag” Cell, 2001, submitted to Nature Structural Biology 2002.

Steven L. Alam*, Owen W. Pornillos*, Rebecca L. Rich, David G. Myszka, Darrell R. Davis, Wesley I. Sundquist ”Structure and functional interactions of the Tsg101 ubiquitin UEV domain” EMBO J. 21(10) 2397-2406.

Zhaoyang Feng, Mark C. Butler, Steven L. Alam and Stewart N. Loh “On the Nature of Conformational Openings: Native- and Unfolded-State Hydrogen and Thiol-Disulfide Exchange Studies of Ferric Aquomyoglobin"” (2001) JMB 314(1) 153-166.

S. L. ALAM, J.F. Atkins, and R.G. Gesteland “Commentary: Programmed Ribosomal Frameshifting: Much ado about knotting” (1999) PNAS,96(25), 14177-14179.

S. L. ALAM, N.M. Wills, J.A. Ingram, J.F. Atkins, and R.G. Gesteland. "Structural Studies of the RNA Pseudoknot Required for Readthrough of the gag-Termination Codon of Murine Leukemia Virus” (1999) JMB, 288, 837-852.