The acrosome is a unique organelle that plays an important role at the site of sperm-zona pellucida binding during the fertilization process, and is lost in globozoospermia, an inherited infertility syndrome in humans. Although the acrosome is known to be derived from the Golgi apparatus, molecular mechanisms underlying acrosome formation are largely unknown. Here we show that Golgi-associated PDZ-and coiled-coil motif-containing protein (GOPC), a recently identified Golgi-associated protein, is predominantly localized at the trans-Golgi region in round spermatids, and male mice in which GOPC has been disrupted are infertile with globozoospermia. The primary defect was the fragmentation of acrosomes in early round spermatids, and abnormal vesicles that failed to fuse to developing acrosomes were apparent. In later stages, nuclear malformation and an abnormal arrangement of mitochondria, which are also characteristic features of human globozoospermia, were observed. Interestingly, intracytoplasmic sperm injection (ICSI) of such malformed sperm into oocytes resulted in cleavage into blastocysts only when injected oocytes were activated. Thus, GOPC provides important clues to understanding the mechanisms underlying spermatogenesis, and the GOPC-deficient mouse may be a unique and valuable model for human globozoospermia.

Abstract. In this paper, we propose a noninvasive method for monitoring threedimensional (3-D) spatial and temporal variations of soil water content in the field, soil moisture tomography. The basic idea of the method originates from Archie's relationship between soil resistivity and water content. Initially, 88 electrodes were densely buried within a 3.5 rn x 3.5 rn square area, and potentials at the electrodes were measured by pole-pole and Wenner array methods at given time intervals. An inversion calculation of the 3-D soil resistivity was then conducted based on these potential data. Next, 46 soil samples were taken at representative positions in the square, and the parameters in the Archie's relationship were measured in the laboratory. Then, the 3-D distributions of the parameters were obtained by a distance weight interpolation method.

Heme-regulated eukaryotic initiation factor 2␣ kinase (HRI) regulates the synthesis of hemoglobin in reticulocytes in response to heme availability. HRI contains a tightly bound heme at the N-terminal domain. Earlier reports show that nitric oxide (NO) regulates HRI catalysis. However, the mechanism of this process remains unclear. In the present study, we utilize in vitro kinase assays, optical absorption, electron spin resonance (ESR), and resonance Raman spectra of purified full-length HRI for the first time to elucidate the regulation mechanism of NO. HRI was activated via heme upon NO binding, and the

Time-resolved UV resonance Raman (UVRR) spectroscopic studies of WT and mutant myoglobin were performed to reveal the dynamics of protein motion after ligand dissociation. After dissociation of carbon monoxide (CO) from the heme, UVRR bands of Tyr showed a decrease in intensity with a time constant of 2 ps. The intensity decrease was followed by intensity recovery with a time constant of 8 ps. On the other hand, UVRR bands of Trp residues located in the A helix showed an intensity decrease that was completed within the instrument response time. The intensity decrease was followed by an intensity recovery with a time constant of Ϸ50 ps and lasted up to 1 ns. The time-resolved UVRR study of the myoglobin mutants demonstrated that the hydrophobicity of environments around Trp-14 decreased, whereas that around Trp-7 barely changed in the primary protein response. The present data indicate that displacement of the E helix toward the heme occurs within the instrument response time and that movement of the FG corner takes place with a time constant of 2 ps. The finding that the instantaneous motion of the E helix strongly suggests a mechanism in which protein structural changes are propagated from the heme to the A helix through the E helix motion.hemeprotein ͉ protein dynamics ͉ resonance Raman spectroscopy ͉ time-resolved spectroscopy P roteins are endowed with both stiff and flexible properties; hence, their dynamics are closely associated with structure and function. Because allosteric proteins, in general, propagate conformational changes over considerable distances, how these conformational changes are generated and transmitted is of major interest for understanding the regulatory, kinetic, and recognition properties of proteins (1-3). A variety of experimental evidence suggests that rapid and long-range propagation of conformational changes through the core of protein plays a vital role in allosteric communication. For example, the cooperative oxygen-binding properties of hemoglobin (Hb) result from a change in quaternary structure, which is initiated by ligand binding/release at the heme (ligand binding site). Therefore, if the pathway by which one quaternary structure is converted to the other quaternary structure is structurally characterized, our understanding how a protein performs its function will be greatly advanced. The ligand-induced dynamics of myoglobin (Mb) are a basic subject for studying such features in proteins. Although Mb is a monomeric protein, the threedimensional structure of Mb is closely similar to that of a subunit of Hb. Thus, the structural changes of Mb can be regarded as a model for the tertiary structural events that cause the quaternary structural change of Hb.

The heme environments of Met 95 and His 77 mutants of the isolated heme-bound PAS domain (Escherichia coli DOS PAS) of a direct oxygen sensing protein from E. coli (E. coli DOS) were investigated with resonance Raman (RR) spectroscopy and compared with the wild type (WT) enzyme. The RR spectra of both the reduced and oxidized WT enzyme were characteristic of six-coordinate low spin heme complexes from pH 4 to 10. The time-resolved RR spectra of the photodissociated CO-WT complex had an iron-His stretching band ( Fe-His ) at 214 cm ؊1 , and the Fe-CO versus CO plot of CO-WT E. coli DOS PAS fell on the line of His-coordinated heme proteins. The photodissociated CO-H77A mutant complex did not yield the Fe-His band but gave a Fe-Im band in the presence of imidazole. The RR spectrum of the oxidized M95A mutant was that of a six-coordinate low spin complex (i.e. the same as that of the WT enzyme), whereas the reduced mutant appeared to contain a fivecoordinate heme complex. Taken Heme-containing signal-transducing proteins (1-3) respond to diatomic molecules, which act as physiological, environmental messengers. This has attracted the attention of biophysical chemists. The O 2 sensing proteins so far identified include FixL (an oxygen-sensing kinase of Rhizobia meliloti) (1, 4), HemAT (an oxygen sensor heme protein discovered from Bacillus subtilis (HemAT-Bs) and Halobacterium salinarium (HemAT-Hs)) (5, 6), PDEA1 (7), and putatively a heme protein from E. coli (designated Escherichia coli DOS) (8). There is only one CO sensor protein known (CooA, a CO-binding transcriptional regulation factor from Rhodospirillum rubrum) (9, 10) and one NO sensor (soluble guanylate cyclase) (11,12). In each case, binding of an external ligand to the heme located in an N-terminal sensory domain transmits a signal to the functional C-terminal domain (either enzymatic or DNA binding). We are curious to know how these proteins recognize a specific diatomic molecule to generate the appropriate physiological response and what kind of structural changes occur to transmit the signal from the sensory domain to the functional domain.The sensory domain of FixL belongs to the large family of signal-transducing PAS domain 1 proteins, whereas those of HemAT, CooA, and soluble guanylate cyclase do not. The PAS domain proteins found in eukarya, archaea, and bacteria contain a partly conserved tertiary structure despite their limited sequence homology (Ͻ15%) and dissimilar cofactors (13). Although structures of three PAS proteins including the human voltage sensor (HERG) (14), the rhizobial oxygen sensor (FixL) (15, 16), and bacterial light sensor (PYP) (17) have been solved, interactions between the sensory domain and the functional domain are not clearly understood. Namely, hydrophobic interactions seem important to regulate the K ϩ channel of HERG, whereas polar interactions in the EF loop of the PAS domain seem to be essential to PYP. In the case of FixL, either a protein conformational change associated with the location of the heme iron (in-pla...