title: Crystal structure of the Pyrococcus horikoshii isopropylmalate isomerase small subunit provides insight into the dual substrate specificity of the enzyme.
abstract: Recent studies have implied that the isopropylmalate isomerase small subunit of the hyperthermophilic archaea Pyrococcus horikoshii (PhIPMI-s) functions as isopropylmalate isomerase in the leucine biosynthesis pathway, and as homoaconitase (HACN) in the lysine biosynthesis pathway via alpha-aminoadipic acid. PhIPMI is thus considered a key to understanding the fundamental metabolism of the earliest organisms. We describe for the first time the crystal structure of PhIPMI-s, which displays dual substrate specificity. The crystal structure unexpectedly shows that four molecules create an interlocked assembly with intermolecular disulfide linkages having a skewed 222 point-group symmetry. Although the overall fold of the PhIPMI-s monomer is related closely to domain 4 of the aconitase (ACN), one alpha-helix in the ACN structure is replaced by a short loop with relatively high temperature factor values. Because this region is essential for discriminating the structurally similar substrate based on interactions with its diversified gamma-moiety, the loop structure in the PhIPMI-s must be dependent on the presence of a substrate. The flexibility of the loop region might be a structural basis for recognizing both hydrophobic and hydrophilic gamma-moieties of two distinct substrates, isopropylmalate and homocitrate.
authors.affiliation: Division of Biological Sciences, Graduate School of Science, Hokkaido University, Kita-10, Nishi-8, Kita-ku, Sapporo, Hokkaido 060-0810, Japan.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Yasutake
authors.forename: Min
authors.initials: M
authors.lastname: Yao
authors.forename: Naoki
authors.initials: N
authors.lastname: Sakai
authors.forename: Tomomi
authors.initials: T
authors.lastname: Kirita
authors.forename: Isao
authors.initials: I
authors.lastname: Tanaka
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000737:chemistry; D019143:Evolution, Molecular; D006836:Hydro-Lyases / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D007535:Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007930:Leucine / Q000096:biosynthesis; D008239:Lysine / Q000096:biosynthesis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D043902:Pyrococcus horikoshii / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D019843:Archaeal Proteins; D004220:Disulfides; D021122:Protein Subunits; D006836:Hydro-Lyases; C032241:isopropylmalate isomerase; C107855:homoaconitate hydratase; D007535:Isomerases; D007930:Leucine; D008239:Lysine