Spatiotemporal brain activity in mental rotation.

There are many higher-order cognitive processes whose neural mechanisms are hard to study by using nonhuman primates. The mental rotation task is one of the best studied of these. We investigated the spatiotemporal brain activity underlying mental rotation of different kinds of stimuli by measuring the magnetoencephalogram of subjects performing two kinds of mental rotation tasks. Visual stimuli in one experiment consisted of hand shapes presented at various orientations, and those in the other consisted of a set of alphabetic characters and their mirror images presented at various orientations. All stimuli were presented in the left visual field. Activity associated with visual stimulus processing was estimated in the lateral occipital lobe, basal occipitotemporal area, and inferior temporal gyrus. Activities related to higher visual processing were differed between the two kinds of stimuli we used. For the hand shape experiment, we found right inferior parietal lobule activity at 200-300 ms after the stimulus presentation. For the alphabetic characters, activity was found in the left superior temporal region at about 300 ms. Furthermore, activity related to mental stimulation in the inferior parietal lobule and premotor area were seen in the both experiments. In the experiments with hands, the premotor activity showed left-hemispheric dominance. However, in the experiments with alphabetic characters, the laterality of the premotor activity did not show left-hemispheric dominance. We inferred this difference was influenced by the subjects' familiarity with the mental rotation of visual stimuli.

Isolation and cDNA-derived amino acid sequences of hemoglobin and myoglobin from the deep-sea clam Calyptogena kaikoi.

The heterodont clam Calyptogena kaikoi, living in the cold-seep area at a depth of 3761 m of the Nankai Trough, Japan, has abundant hemoglobins and myoglobins in erythrocytes and adductor muscle, respectively. Two types of hemoglobins (Hb I and Hb II) were isolated, and the complete amino acid sequences of Hb I (145 residues) and Hb II (137 residues) were obtained with combination of cDNA and protein sequencing. The amino acid sequences of C. kaikoi Hbs I and II differed from homologous chains of the congeneric clam Calyptogena soyoae in eight and five positions, respectively. The distal (E7) His, one of the functionally important residues in hemoglobin and myoglobin, was replaced by Gln in hemoglobins of C. kaikoi. A phylogenetic analysis of clam hemoglobins indicates that the evolutionary rate of Calyptogena hemoglobins is rather faster than those of other clams, suggesting that the mutation rate might be accelerated in the deep-sea animals around the areas of cold seeps or hydrothermal vents. On the other hand, it was found unexpectedly that two myoglobins Mbs I and II, isolated from the red adductor muscle, are identical in amino acid sequence Hbs I and II, respectively. Thus it was assumed that genes for Hbs I and II are also expressed in the muscle of C. kaikoi in substitution for myoglobin gene. This suggests that the major physiological role of globins in C. kaikoi is storage of oxygen under the low oxygen conditions, rather than circulating of oxygen.

Arginine kinase evolved twice: evidence that echinoderm arginine kinase originated from creatine kinase.

Arginine kinase (AK) was isolated from the longitudinal muscle of the sea cucumber Stichopus japonicus. Unlike the monomeric 40 kDa AKs from molluscs and arthropods, but like the cytoplasmic isoenzymes of vertebrate creatine kinase (CK), the Stichopus enzyme was dimeric. To explore the evolutionary origin of the dimeric AK, we determined its cDNA-derived amino acid sequence of 370 residues. A comparison of the sequence with those of other enzymes belonging to the phosphagen kinase family indicated that the entire amino acid sequence of Stichopus AK is apparently much more similar to vertebrate CKs than to all other AKs. A phylogenetic tree also strongly suggests that the Stichopus AK has evolved from CK. These results support the conclusion that AK evolved at least twice during the evolution of phosphagen kinases: first at an early stage of phosphagen kinase evolution (its descendants are molluscan and arthropod AKs) and secondly from CK later in metazoan evolution. A comparison of the amino acid sequence around the guanidino specificity (GS) region (which is a possible candidate for the guanidine substrate recognition site in the phosphagen kinase family) of the Stichopus enzyme with those of other phosphagen kinases showed that the GS region of the Stichopus enzyme was of the AK type: five amino acid deletions in the flexible loop region that might help to accommodate larger guanidine substrates in the active site. The presence of the AK-type deletions in the Stichopus AK, even though it seems that the enzyme's most immediate ancestor was probably CK, strongly suggests that the GS region has a role in substrate specificity. Stichopus AK and presumably other echinoderm AKs seem to have evolved from the CK gene; the sequence of GS region might have been replaced by the AK type via exon shuffling. The presence of an intron near the GS region in the Stichopus AK gene supports this hypothesis.

Evolution of phosphagen kinase VII. Isolation of glycocyamine kinase from the polychaete Neanthes diversicolor and the cDNA-derived amino acid sequences of alpha and beta chains.

Glycocyamine kinase (GK) was isolated from the marine polychaete Neanthes diversicolor by gel filtration, DEAE-cellulose chromatography, butyl-Toyopearl hydrophobic chromatography, and chromatofocusing. The GK was eluted as a single peak on the latter three chromatographies, and the molecular mass for the native GK was estimated to be about 80 kDa. The SDS-PAGE showed that the isolated GK consists of two distinct subunits in equal proportion, alpha and beta chains, with molecular masses of 42.2 and 43.8 kDa, respectively. The present results suggest that the Neanthes GK has a heterodimeric structure. The cDNAs for alpha and beta chains of Neanthes GK were amplified by PCR and their cDNA-derived amino acid sequences were determined. The alpha and beta chains are composed of 374 and 390 amino acids, and the molecular masses were calculated to be 42,392 and 43,966 Da, respectively, in good agreement with the apparent masses on SDS PAGE. The beta chain has a characteristic N-terminal extension of 15 amino acids, and all of the sequence differences between alpha and beta chains were restricted in the N-terminal region of 50 residues. The overall sequence identity was 92%. The occurrence of heterodimeric nature in Neanthes GK is of great interest from the evolutionary point of view, because the heterodimeric structure is only known for creatine kinase MB-isozyme specific for mammalian heart muscle among phosphagen kinases.

The cDNA-derived amino acid sequence of indoleamine dioxygenase like-myoglobin from the gastropod mollusc Omphalius pfeifferi.

Myoglobin was isolated from the radular muscle of the archaegastropod mollusc Omphalius pfeifferi (Trochidae). The molecular mass was estimated by SDS-PAGE to be about 40 kDa, 2.5 times larger than that of usual myoglobin. The cDNA for Omphalius myoglobin was amplified by polymerase chain reaction, and the cDNA-derived amino acid sequence of 375 residues was determined, of which 73 residues were identified directly by the chemical sequencing of internal peptides. The amino acid sequence of Omphalius myoglobin showed no significant homology with any other usual 16-kDa globins, but showed 84% and 36% identities with indoleamine dioxygenase-like myoglobins from Battilus (Turbinidae) and Sulculus (Haliotiidae), respectively. It also shows significant homology (26% identity) with human indoleamine 2,3-dioxygenase, a tryptophan-degrading enzyme containing heme. The distribution of indoleamine dioxygenase-like myoglobins suggests that they must have arisen exclusively along the specified lineage including the three families Haliotiidae, Turbinidae, and Trochidae of Archaegastropoda in molluscan evolution.

Temporal structure of implicit motor imagery in visual hand-shape discrimination as revealed by MEG.

We investigated the spatio-temporal brain activity on the time scale of several milliseconds related to the mental rotation task requiring judgements of hand orientation, using a whole-cortex MEG (magnetoencephalography) system. Neuronal activity in the visual cortex was observed approximately 100-200 ms from stimulus onset, and that in inferior parietal lobe followed (after 200 ms). Both of these activities showed a contralateral dominance to visual stimulus hemifield. Premotor activity started later than the inferior parietal lobe activity, and these activities partially overlapped. Activity in primary motor and/or motosensory areas was observed in some subjects. The whole-cortex neuromagnetic measurements provided the time course of activity in the human brain associated with the implicit motor imagery: visual cortex-->inferior parietal lobe<-->premotor cortex. This process is considered to be the transformation process of retinotopic locations into a body-centered reference frame necessary for the mental rotation task.

Amino acid sequence, spectral, oxygen-binding, and autoxidation properties of indoleamine dioxygenase-like myoglobin from the gastropod mollusc Turbo cornutus.

Myoglobin was isolated from the radular muscle of the archaeogastropod mollusc Turbo cornutus (Turbinidae). This myoglobin is a monomer carrying one protoheme group; the molecular mass was estimated by SDS-PAGE to be about 40 kDa, 2.5 times larger than that of usual myoglobin. The cDNA-derived amino acid sequence of 375 residues was determined, of which 327 residues were identified directly by chemical sequencing of internal peptides. The amino acid sequence of Turbo myoglobin showed no significant homology with any other usual 16-kDa globins, but showed 36% identity with the myoglobin from Sulculus diversicolor (Haliotiidae) and 27% identity with human indoleamine 2,3-dioxygenase, a tryptophan-degrading enzyme containing heme. Thus, the Turbo myoglobin can be counted among the myoglobins which evolved from the same ancestor as that of indoleamine 2,3-dioxygenase. The absorbance ratio of gamma to CT maximum (gamma/CT) of Turbo metmyoglobin was 17.8, indicating that this myoglobin probably possesses a histidine residue near the sixth coordination position of heme iron. The Turbo myoglobin binds oxygen reversibly. Its oxygen equilibrium properties are similar to those of Sulculus myoglobin, giving P50 = 3.5 mm Hg at pH 7.4 and 20 degrees C. The pH dependence of autoxidation of Turbo oxymyoglobin was quite different from that of mammalian myoglobin, suggesting a unique protein folding around the heme cavity of Turbo myoglobin. A kinetic analysis of autoxidation indicates that the amino acid residue with pKa = 5.4 is involved in the reaction. The autoxidation reaction was enhanced markedly at pH 7.6, but not at pH 5.5 and 6.3 in the presence of tryptophan. We suggest that a noncatalytic binding site for tryptophan, in which several dissociation groups with pKa > or = 7.6 are involved, remains in Turbo myoglobin as a relic of molecular evolution.

A myoglobin evolved from indoleamine 2,3-dioxygenase, a tryptophan-degrading enzyme.

The distribution, isolation, spectral and oxygen-binding properties, stability of ferrous state (autoxidation), amino acid sequence and gene structure of indoleamine 2,3-dioxygenase (IDO)-like myoglobins are summarized, and their evolution is discussed. Although it has long been thought that all hemoglobins and myoglobins have evolved from a common ancestral gene encoding a 14-16 kDa polypeptide, the discovery of IDO-like myoglobin from several gastropod molluscs clearly indicates that there was an alternative pathway for myoglobin evolution.