The Helix-Loop-Helix (HLH) family of eukaryotic transcription factors comprises a large number of proteins which play key roles in homeostasis, the regulation cell proliferation, and differentiation. These proteins share a phylogenetically conserved bipartite b/HLH domain responsible for specific DNA binding and dimerization. The HLH region dictates dimerization affinity and specificity while the basic region (b) is primarily responsible for sequence-specific DNA binding. In some family members, such as the Myc oncoproteins, the HLH motif is followed by a heptad repeat of hydrophobic amino acids, or \"leucine zipper\" (Z). My X-ray crystallographic structure determination at 2.9Å resolution of a dimer of the b/HLH/Z domain of the mammalian oncoprotein Max bound to its target DNA revealed that this symmetric homodimer folds into a novel parallel left-handed four-helix bundle, which is globular and stabilized by a well-defined hydrophobic core. Two pairs of α-helices protrude in opposite directions from the bundle. One, the basic regions, enters the major groove of the target B-form DNA and makes numerous contacts with the bases and phosphodiester backbone. The other, the leucine zipper, forms a left-handed coiled-coil, extending the hydrophobic interface of the homodimer. I also determined the cocrystal structure of a truncated b/HLH homodimer of the human transcription factor USF bound to DNA. As expected from the sequence conservation, this protein adopts the same three-dimensional structure as Max b/HLH. Circular dichroism spectroscopic investigation of DNA binding by Max and USF demonstrated, in concert with their cocrystal structures, that these proteins undergo a dramatic folding transition upon specific, high-affinity DNA binding. More than forty residues per dimer become α-helical upon association. I also demonstrated by hydrodynamic as well as biochemical methods that these proteins can form bivalent tetramers at physiologically meaningful concentrations. This suggests that they may play a role in DNA looping, thought to be important in the transcriptional regulation of eukaryotic genes.
","abstract_html":"<p>The Helix-Loop-Helix (HLH) family of eukaryotic transcription factors comprises a large number of proteins which play key roles in homeostasis, the regulation cell proliferation, and differentiation. These proteins share a phylogenetically conserved bipartite b/HLH domain responsible for specific DNA binding and dimerization. The HLH region dictates dimerization affinity and specificity while the basic region (b) is primarily responsible for sequence-specific DNA binding. In some family members, such as the Myc oncoproteins, the HLH motif is followed by a heptad repeat of hydrophobic amino acids, or "leucine zipper" (Z). My X-ray crystallographic structure determination at 2.9Å resolution of a dimer of the b/HLH/Z domain of the mammalian oncoprotein Max bound to its target DNA revealed that this symmetric homodimer folds into a novel parallel left-handed four-helix bundle, which is globular and stabilized by a well-defined hydrophobic core. Two pairs of α-helices protrude in opposite directions from the bundle. One, the basic regions, enters the major groove of the target B-form DNA and makes numerous contacts with the bases and phosphodiester backbone. The other, the leucine zipper, forms a left-handed coiled-coil, extending the hydrophobic interface of the homodimer. I also determined the cocrystal structure of a truncated b/HLH homodimer of the human transcription factor USF bound to DNA. As expected from the sequence conservation, this protein adopts the same three-dimensional structure as Max b/HLH. Circular dichroism spectroscopic investigation of DNA binding by Max and USF demonstrated, in concert with their cocrystal structures, that these proteins undergo a dramatic folding transition upon specific, high-affinity DNA binding. More than forty residues per dimer become α-helical upon association. I also demonstrated by hydrodynamic as well as biochemical methods that these proteins can form bivalent tetramers at physiologically meaningful concentrations. This suggests that they may play a role in DNA looping, thought to be important in the transcriptional regulation of eukaryotic genes.</p>","abstract_has_math":false,"creators":["Ferre-D'Amare, Adrian"],"institution":null,"degree_name":"Doctor of Philosophy (PhD)","degree_level":"Thesis","degree_discipline":null,"degree_department":null,"school":null,"contributors":["Stephen Burley"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1995,"date_issued":"1995-01-01T08:00:00Z","date_published":"1995-01-01T08:00:00Z","updated_at":"2026-07-24T04:11:45Z","subjects":["HLH transcription factors","DNA binding","Max protein","USF structure","protein dimerization","X-ray crystallography","Life Sciences"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/620","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Stephen Burley"]},{"key":"dc:creator","label":"Author","values":["Ferre-D'Amare, Adrian"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"thesis:degree_level","label":"Degree Level","values":["Thesis"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Doctor of Philosophy (PhD)"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["HLH transcription factors","DNA binding","Max protein","USF structure","protein dimerization","X-ray crystallography","Life Sciences"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/620"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The Helix-Loop-Helix (HLH) family of eukaryotic transcription factors comprises a large number of proteins which play key roles in homeostasis, the regulation cell proliferation, and differentiation. These proteins share a phylogenetically conserved bipartite b/HLH domain responsible for specific DNA binding and dimerization. The HLH region dictates dimerization affinity and specificity while the basic region (b) is primarily responsible for sequence-specific DNA binding. In some family members, such as the Myc oncoproteins, the HLH motif is followed by a heptad repeat of hydrophobic amino acids, or \"leucine zipper\" (Z). My X-ray crystallographic structure determination at 2.9Å resolution of a dimer of the b/HLH/Z domain of the mammalian oncoprotein Max bound to its target DNA revealed that this symmetric homodimer folds into a novel parallel left-handed four-helix bundle, which is globular and stabilized by a well-defined hydrophobic core. Two pairs of α-helices protrude in opposite directions from the bundle. One, the basic regions, enters the major groove of the target B-form DNA and makes numerous contacts with the bases and phosphodiester backbone. The other, the leucine zipper, forms a left-handed coiled-coil, extending the hydrophobic interface of the homodimer. I also determined the cocrystal structure of a truncated b/HLH homodimer of the human transcription factor USF bound to DNA. As expected from the sequence conservation, this protein adopts the same three-dimensional structure as Max b/HLH. Circular dichroism spectroscopic investigation of DNA binding by Max and USF demonstrated, in concert with their cocrystal structures, that these proteins undergo a dramatic folding transition upon specific, high-affinity DNA binding. More than forty residues per dimer become α-helical upon association. I also demonstrated by hydrodynamic as well as biochemical methods that these proteins can form bivalent tetramers at physiologically meaningful concentrations. This suggests that they may play a role in DNA looping, thought to be important in the transcriptional regulation of eukaryotic genes.
"]},{"key":"dc:title","label":"Title","values":["DNA Recognition by Helix-Loop-Helix Proteins"]}]}],"canonical_facts":{"dc:contributor":["Stephen Burley"],"dc:creator":["Ferre-D'Amare, Adrian"],"dc:description.abstract":["The Helix-Loop-Helix (HLH) family of eukaryotic transcription factors comprises a large number of proteins which play key roles in homeostasis, the regulation cell proliferation, and differentiation. These proteins share a phylogenetically conserved bipartite b/HLH domain responsible for specific DNA binding and dimerization. The HLH region dictates dimerization affinity and specificity while the basic region (b) is primarily responsible for sequence-specific DNA binding. In some family members, such as the Myc oncoproteins, the HLH motif is followed by a heptad repeat of hydrophobic amino acids, or \"leucine zipper\" (Z). My X-ray crystallographic structure determination at 2.9Å resolution of a dimer of the b/HLH/Z domain of the mammalian oncoprotein Max bound to its target DNA revealed that this symmetric homodimer folds into a novel parallel left-handed four-helix bundle, which is globular and stabilized by a well-defined hydrophobic core. Two pairs of α-helices protrude in opposite directions from the bundle. One, the basic regions, enters the major groove of the target B-form DNA and makes numerous contacts with the bases and phosphodiester backbone. The other, the leucine zipper, forms a left-handed coiled-coil, extending the hydrophobic interface of the homodimer. I also determined the cocrystal structure of a truncated b/HLH homodimer of the human transcription factor USF bound to DNA. As expected from the sequence conservation, this protein adopts the same three-dimensional structure as Max b/HLH. Circular dichroism spectroscopic investigation of DNA binding by Max and USF demonstrated, in concert with their cocrystal structures, that these proteins undergo a dramatic folding transition upon specific, high-affinity DNA binding. More than forty residues per dimer become α-helical upon association. I also demonstrated by hydrodynamic as well as biochemical methods that these proteins can form bivalent tetramers at physiologically meaningful concentrations. This suggests that they may play a role in DNA looping, thought to be important in the transcriptional regulation of eukaryotic genes.
"],"dc:identifier":["https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/620"],"dc:subject":["HLH transcription factors","DNA binding","Max protein","USF structure","protein dimerization","X-ray crystallography","Life Sciences"],"dc:title":["DNA Recognition by Helix-Loop-Helix Proteins"],"thesis:degree_level":["Thesis"],"thesis:degree_name":["Doctor of Philosophy (PhD)"]},"updated_at":"2026-07-24T04:11:45Z"}