Search

A practical method to evaluate the contributions of the nonlinear polarizability and anharmonicity of potentials from the experimental and simulation data by using double Fourier transformation is presented. In a Lennard-Jones potential system, an approximated expression of the fifth-order response function using the ratio between nonlinear polarizability and anharmonicity exhibits a good agreement with the results of the molecular dynamics simulation. In a soft-core case, the fifth-order Raman signal indicates that the system consists of the delocalized and localized modes, and only the delocalized mode affects the dramatic change of the fifth-order Raman response functions between solid and liquid phases through nonlinear polarizability.

Gamma-secretase cleaves the transmembrane domain of beta-amyloid precursor protein at multiple sites. These are referred to as gamma-, zeta-, and epsilon-cleavages. We showed previously that DAPT, a potent dipeptide gamma-secretase inhibitor, caused differential accumulations of longer amyloid beta-proteins (Abetas) (Abeta43 and Abeta46) in CHO cells that are induced to express the beta C-terminal fragment (CTF). To learn more about the cleavage mechanism by gamma-secretase, CHO cell lines coexpressing betaCTF and wild-type or mutant presenilin (PS) 1/2 were generated and treated with DAPT. In all cell lines treated with DAPT, as the levels of Abeta40 decreased, Abeta46 accumulated to varying extents. In wild-type PS1 or M146L mutant PS1 cells, substantial amounts of Abeta43 and Abeta46 accumulated. In contrast, this was not the case with wild-type PS2 cells. In M233T mutant PS1 cells, significant amounts of Abeta46 and Abeta48 accumulated differentially, whereas in N141I mutant PS2 cells, large amounts of Abeta45 accumulated concomitantly with a large decrease in Abeta42 levels. Most interestingly, in G384A mutant PS1 cells, there were no significant accumulations of longer Abetas except for Abeta46. Abeta40 was very susceptible to DAPT, but other Abetas were variably resistant. Complicated suppression and accumulation patterns by DAPT may be explained by stepwise processing of betaCTF from a zeta- or epsilon-cleavage site to a gamma-cleavage site and its preferential suppression of gamma-cleavage over zeta- or epsilon-cleavage.

Ionic solvation in a polar solvent is modeled by a central charge surrounded by dipolar molecules posted on two-dimensional distorted lattice sites with simple rotational dynamics. Density of states is calculated by applying the Wang-Landau algorithm to both the energy and polarization states. The free energy landscapes of solvent molecules as a function of polarization are depicted to explore the competition between the thermal fluctuation and solvation energy. Without a central charge, for temperatures higher than the energy scale of the dipole-dipole interactions, the energy landscape for the small polarization region exhibits a parabolic shape as predicted by Marcus [Rev. Mod. Phys. 65, 599 (1993)] for electron transfer reaction, while there is an additional quartic contribution to the landscape for the large polarization region. When the temperature drops, the simulated free energy landscapes are no longer smooth due to the presence of multiple local minima arising from the frustrated interaction among the dipoles. The parabolic contribution becomes negligible and the energy landscape becomes quartic in shape. For a strong central charge, the energy landscape exhibits an asymmetric profile due to the contributions of linear and cubic terms that arise from the charge-dipole interactions.

We calculate third- and fifth-order Raman spectra of simple atoms interacting through a soft-core potential by means of molecular-dynamics (MD) simulations. The total polarizability of molecules is treated by the dipole-induced dipole model. Two- and three-body correlation functions of the polarizability at various temperatures are evaluated from equilibrium MD simulations based on a stability matrix formulation. To analyze the processes involved in the spectroscopic measurements, we divide the fifth-order response functions into symmetric and antisymmetric integrated response functions; the symmetric one is written as a simple three-body correlation function, while the antisymmetric one depends on a stability matrix. This analysis leads to a better understanding of the time scales and molecular motions that govern the two-dimensional (2D) signal. The 2D Raman spectra show novel differences between the solid and liquid phases, which are associated with the decay rates of coherent motions. On the other hand, these differences are not observed in the linear Raman spectra.

We present an analytical expression for the linear and nonlinear infrared spectra of interacting molecular vibrational motions. Each of the molecular modes is explicitly represented by a classical damped oscillator on an anharmonic multidimensional potential-energy surface. The two essential interactions, the dipole-dipole (DD) and the dipole-induced-dipole (DID) interactions, are taken into account, and each dipole moment and polarizability are expanded to nonlinear order with respect to the nuclear vibrational coordinate. Our analytical treatment leads to expressions for the contributions of anharmonicity, DD and DID interactions, and the nonlinearity of dipole moments and polarizability elements to the one-, two-, and three-dimensional spectra as separated terms, which allows us to discuss the relative importance of these respective contributions. We can calculate multidimensional signals for various configurations of molecules interacting through DD and DID interactions for different material parameters over the whole range of frequencies. We demonstrate that contributions from the DD and DID interactions and anharmonicity are separately detectable through the third-order three-dimensional IR spectroscopy, whereas they cannot be distinguished from each other in either the linear or the second-order IR spectroscopies. The possibility of obtaining the intra- or intermolecular structural information from multidimensional spectra is also discussed.

Simulating tunneling processes as well as their observation are challenging problems for many areas. In this study, we consider a double-well potential system coupled to a heat bath with a linear-linear (LL) and square-linear (SL) system-bath interactions. The LL interaction leads to longitudinal (T1) and transversal (T2) homogeneous relaxations, whereas the SL interaction leads to the inhomogeneous dephasing (T2*) relaxation in the white noise limit with a rotating wave approximation. We discuss the dynamics of the double-well system under infrared (IR) laser excitations from a Gaussian-Markovian quantum Fokker-Planck equation approach, which was developed by generalizing Kubo's stochastic Liouville equation. Analytical expression of the Green function is obtained for a case of two-state-jump modulation by performing the Fourier-Laplace transformation. We then calculate a two-dimensional infrared signal, which is defined by the four-body correlation function of optical dipole, for various noise correlation time, system-bath coupling parameters, and temperatures. It is shown that the bath-induced vibrational excitation and relaxation dynamics between the tunneling splitting levels can be detected as the isolated off-diagonal peaks in the third-order two-dimensional infrared (2D-IR) spectroscopy for a specific phase matching condition. Furthermore, this spectroscopy also allows us to directly evaluate the rate constants for tunneling reactions, which relates to the coherence between the splitting levels; it can be regarded as a novel technique for measuring chemical reaction rates. We depict the change of reaction rates as a function of system-bath coupling strength and a temperature through the 2D-IR signal.

To examine how gamma- and epsilon-cleavages of beta-amyloid precursor protein (APP) are related, each cleavage site was replaced with a stretch of Trp that cannot be cleaved by gamma-secretase. Replacement of the gamma- or epsilon-site significantly suppressed secretion of amyloid beta-protein (Abeta), and produced longer Abeta or longer APP intracellular domain, respectively. This cleavage at the midportion between gamma- and epsilon-sites was also gamma-secretase-dependent. Blocking this cleavage with a Trp stretch remarkably suppressed Abeta generation, indicating that the midportion cleavage is required for the generation of Abeta.

We studied the effect of TNF-alpha stimulation on a cholangiocarcinoma cell line, CCKS1. CCKS1 expressed only one type TNF receptor, TNFR2. Treatment of CCKS1 with TNF-alpha substantially activated NFkappaB, MAPK and Akt signalings which in turn activated matrix metalloproteinase-9 (MMP-9) secretion and in vitro invasiveness of CCKS1. Pretreatment of cells with anti-TNFR2 neutralizing antibody inhibited the TNF-alpha-dependent signaling and MMP-9 secretion and subsequently blocked invasion in vitro. Moreover, an inhibitor for matrix metalloproteinase, Galardin, suppressed the invasion in a dose-dependent manner. Similarly, pharmacological inhibition of signaling clearly suppressed the TNF-alpha dependent MMP-9 secretion. These results strongly suggest that TNF-alpha-TNFR2 signaling plays an important role to convert the cholangiocarcinoma cells to be more aggressive one.

Gamma-cleavage of beta-amyloid precursor protein (APP) in the middle of the cell membrane generates amyloid beta protein (Abeta), and epsilon-cleavage, approximately 10 residues downstream of the gamma-cleavage site, releases the APP intracellular domain (AICD). A significant link between generation of Abeta and AICD and failure to detect AICD41-99 led us to hypothesize that epsilon-cleavage generates longer Abetas, which are then processed to Abeta40/42. Using newly developed gel systems and an N-end-specific monoclonal antibody, we have identified the longer Abetas (Abeta1-43, Abeta1-45, Abeta1-46, and Abeta1-48) within the cells and in brain tissues. The production of these longer Abetas as well as Abeta40/42 is presenilin dependent and is suppressed by {1S-benzyl-4R-[1S-carbamoyl-2-phenylethylcarbamoyl-1S-3-methylbutylcarbamoyl]-2R-hydroxy-5-phenylpentyl}carbamic acid tert-butyl ester, a transition state analog inhibitor for aspartyl protease. In contrast, N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester, a potent dipeptide gamma-secretase inhibitor, builds up Abeta1-43 and Abeta1-46 intracellularly, which was also confirmed by mass spectrometry. Notably, suppression of Abeta40 appeared to lead to an increase in Abeta43, which in turn brings an increase in Abeta46, in a dose-dependent manner. We therefore propose an alpha-helical model in which longer Abeta species generated by epsilon-cleavage is cleaved at every three residues in its carboxyl portion.

We previously showed that beta-amyloid precursor protein (APP) is cleaved not only in the middle of the membrane (gamma-cleavage) but also at novel cleavage sites close to the membrane/cytoplasmic boundary (epsilon-cleavage), releasing APP intracellular domains (AICDs) 49-99 and 50-99. To learn more about the relationship between gamma- and epsilon-cleavage, C-terminally truncated carboxyl-terminal fragments (CTFs) of APP, especially CTFs1-48 and 1-49 (the postulated products that are generated by epsilon-cleavage), were transiently expressed in CHO cells. Most importantly, the cells expressing CTF1-49 secreted predominantly amyloid beta-protein (Abeta) 40, while those expressing CTF1-48 secreted preferentially Abeta42. This supports our assumption that epsilon-cleavage precedes Alphabeta production and that preceding epsilon-cleavage determines the preference for the final Abeta species. The gamma-secretase inhibitors, L-685,458 and DAPT, suppressed Abeta production from CTF1-49. Regarding Abeta production from CTF1-48, L-685,458 suppressed it, but DAPT failed to do so. A dominant negative mutant of presenilin 1 suppressed the production of Abeta40 and 42 from both CTFs1-48 and 1-49. These data should shed significant light into the mechanism of Abeta production.

Spectrum changeable neutron calibration fields are planned to be established with an accelerator installed in Japan Atomic Energy Research Institute/Facility of Radiation Standards. The neutron fields are provided by bombarding a target surrounded by a moderator, with charged particles from the accelerator. In the fields, a wide variety of neutron spectra is provided with sufficient fluence rate for the calibration of dosemeters. In this study, necessity of the field was first discussed in view of relationship between readings of existing dosemeters and true dose equivalents where the dosemeters were used. Second, test simulation of neutron spectra was carried out with the Monte Carlo technique for some arrangements with a LiF target and quasi-cylindrical moderators with different materials. The simulated spectra were summarised in terms of fluence-average energy, fluence rate and calibration factor for the dosemeters.

Multidimensional vibrational response functions of a harmonic oscillator are reconsidered by assuming nonlinear system-bath couplings. In addition to a standard linear-linear (LL) system-bath interaction, we consider a square-linear (SL) interaction. The LL interaction causes the vibrational energy relaxation, while the SL interaction is mainly responsible for the vibrational phase relaxation. The dynamics of the relevant system are investigated by the numerical integration of the Gaussian-Markovian Fokker-Planck equation under the condition of strong couplings with a colored noise bath, where the conventional perturbative approach cannot be applied. The response functions for the fifth-order nonresonant Raman and the third-order infrared (or equivalently the second-order infrared and the seventh-order nonresonant Raman) spectra are calculated under the various combinations of the LL and the SL coupling strengths. Calculated two-dimensional response functions demonstrate that those spectroscopic techniques are very sensitive to the mechanism of the system-bath couplings and the correlation time of the bath fluctuation. We discuss the primary optical transition pathways involved to elucidate the corresponding spectroscopic features and to relate them to the microscopic sources of the vibrational nonlinearity induced by the system-bath interactions. Optical pathways for the fifth-order Raman spectroscopies from an "anisotropic" medium were newly found in this study, which were not predicted by the weak system-bath coupling theory or the standard Brownian harmonic oscillator model., ((c) 2004 American Institute of Physics)

Monoenergetic neutron calibration fields of 144, 565 keV and 5.0 MeV have been developed at the Facility of Radiation Standards of JAERI using a 4 MV Pelletron accelerator. The 7Li(p,n)7Be and 2H(d,n)3He reactions are employed for neutron production. The neutron energy was measured by the time-of-flight method with a liquid scintillation detector and calculated with the MCNP-ANT code. A long counter is employed as a neutron monitor because of the flat response. The monitor is set up where the influence of inscattered neutrons from devices and their supporting materials at a calibration point is as small as possible. The calibration coefficients from the monitor counts to the neutron fluence at a calibration point were obtained from the reference fluence measured with the transfer instrument of the primary standard laboratory (AIST), a 24.13 cm phi Bonner sphere counter. The traceability of the fields to AIST was established through the calibration.

The Facility of Radiation Standards of Japan Atomic Energy Research Institute has been equipped with concrete-moderated neutron calibration fields as simulated workplace neutron fields. The fields use an 241Am-Be neutron source placed in the narrow space surrounded by concrete bricks, walls and floor. The neutron spectra and the neutron fluence rates of the fields were measured with the Bonner multi-sphere spectrometer system (BMS), spherical recoil-proton proportional counters, and a liquid scintillation counter (NE-213). The results were compared with each other. The reference values of H*(10) were determined from the results of BMS. The angular distributions of neutron fluence were calculated using MCNP-4B2 to obtain the reference values of Hp(10). The calculated results show that the scattered neutrons have a wide range of incident angles. The reference Hp(10) values considered the angular distribution were found to be 10-18% smaller than those without consideration.

The function of the translation product of cDNA for Acetabularia vacuolar H(+)-pyrophosphatase was examined using the Saccharomyces cerevisiae VMA3-deficient strain. The open reading frame of Acetabularia H(+)-pyrophosphatase was revealed to encode 751 amino acids (721 or 751 amino acids in a previous paper). The acidification of the vacuole was observed by fluorescence microscopy when the cDNA was constructed in pYES2. Immunoblot analysis also supported the localization of the translation product in the vacuolar-membrane-enriched fraction.

We compared the cytotoxicity of soluble and particulate titanium (Ti), vanadium (V) and nickel (Ni) by biochemical functional analysis and by microscopic morphology with micro-area elemental analysis in vitro using human neutrophils as probes and in vivo in animals. The biochemical analyses of LDH, superoxide anion, TNF-alpha and scanning electron microscopy (SEM) showed that Ni in solution destroys the cell membrane of neutrophils, whereas Ti and V in solution stimulate neutrophils and increase the quantity of released superoxide anions. Fine Ti particles (1-3 microm), which smaller than neutrophils (about 5 microm), were phagocytized by the cells and the results were similar in vivo. These results showed that the cytotoxic effect of Ti particles is size dependent, and that they must be smaller than that of cells. The present study demonstrated that the biochemical functional tests are useful for evaluating the biocompatibility of materials.

The genes encoding 'cytochrome a1'-like hemoprotein of Magnetospirillum magnetotacticum were identified and sequenced. Three ORFs, mcalI, mcaI and hosA, were included in the sequenced region. The six histidine residues which were predicted to associate with the prosthetic cofactors of heme-copper oxidase superfamily were conserved in the hemoprotein. However, none of the amino acid residues which were proposed to participate in the oxygen-reducing and the coupled proton pumping reactions in cytochrome c oxidase were at all conserved in the hemoprotein.

The topic of femtochemistry is surveyed from both theoretical and experimental points of view. A time-dependent wave packet description of the photodissociation of the O---C---S molecule reveals vibrational motion in the transition-state region and suggests targets for direct experimental observation. Theoretical approaches for treating femtosecond chemical phenomena in condensed phases are featured along with prospects for laser-controlled chemical reactions by using tailored ultrashort chirped pulses. An experimental study of the photoisomerization of retinal in the protein bacteriorhodopsin is discussed with an aim to gain insight into the potential energy surfaces on which this remarkably efficient and selective reactions proceeds. Finally, a prospective view of new frontiers in femtochemistry is given.

The method as well as an appropriate instrumentation for measuring phase changes of reflected light is described. The phase changes on samples of Au, Al, Ag, and Cr evaporated films are measured for five wavelengths (lambda) from 442 to 633 nm, with respect to the phase change at the glass-air interface, where it should be zero. The measured results for the Au film are in fairly good agreement with values calculated by use of optical constants from a handbook or the complex refractive index measured by an ellipsometer. The phase changes for Al and Ag films are different from calculated values by ~5 degrees or a shift length of 4.4 nm at lambda = 633 nm, while those of the Cr film show large shifts as high as 16 degrees or a shift length of 9.8 nm at lambda = 442 nm.

The relationship between Polychlorinated Biphenyls (PCBs), and Polychlorinated Quaterphenyls (PCQs) concentrations in the skin surface lipids and blood was investigated in twenty two patients with PCB poisoning (Yusho) and 9 control subjects were analyzed. The collection of skin surface lipids was performed by the method of cotton pad with 70% ethyl alcohol. The alkaline decomposition method described as "the official standard analytical methods for the isolation of PCBs and PCQs fractions" was used. In the blood of control group, the mean value of PCBs concentration was 1.8 ng/g, and that of PCQs concentration was very low and undetectable by our analytical method. On the other hand, the PCBs concentration in the Yusho group was three times higher than those in the control group. The mean value of PCQs concentration was 1.90 ng/g in Yusho group, while it was not detectable in the control group. In the skin surface lipids of patients with Yusho, the mean value of PCBs concentration was 668.6 ng/g, but was 256.7 ng/g in the control group. The mean value of PCQs concentration in the skin surface lipids of Yusho patients was 35.4 ng/g, although it was not detected in the control group. These values of the skin surface lipids in 1993 were decreased as compared with those in 1992. PCBs and PCQs levels in the skin surface lipids were higher than those in the blood. These findings suggests that cutaneous sebaceous system is one of the excretory systems of polyhalogenated chemicals, such as PCBs or PCQs, when these chemicals are precipitated in human or mammalian.

In this paper, the normal values, distribution, and variations in serum and urinary activities of the lysosomal enzyme, N-acetyl-beta-D-glucosaminidase (NAG), were estimated in a rural area of Japan. The frequency distribution of NAG activities both in the serum and urine of 1,152 males and females (aged 35 to 87), living in a rural area of Gifu Prefecture in Japan, showed an approximately log-normal distribution. Geometric mean values (with S.D. ranges) in healthy subjects (876 males and females) were 12.61 U/l (9.56-16.63) for serum NAG and 8.05 U/l (4.03-16.07) for urinary NAG. There was no significant difference between those of males and females. Serum and urinary NAG activities increased with advancing ages, and these NAG activities in hypertensive subjects were higher than those in normotensives. Since the serum and urinary levels of several commonly measured cytosolic chemicals were within normal limits, the above results could not be ascribed to tissue necrosis. Thus, the estimation of serum and urinary NAG could be a useful indicator discriminating the essential hypertensive patients from the normal subjects.

Plasma concentrations of clioquinol and its metabolites after single or repeated oral administration of clioquinol, absorption region of clioquinol in gastrointestinal tract, and intestinal metabolism were studied in rats. Plasma concentrations of clioquinol after oral administration of four different doses (20, 100, 200 and 400 mg/kg) were lower than those of the two metabolites, clioquinol glucuronide and sulfate. Mean maximal plasma concentration of unchanged drug was in the range of 1-8 nmol/ml. Clioquinol was absorbed poorly from the stomach and fairly from the small intestine. Bile was an important route for excretion of clioquinol in rats. The mesenteric venous plasma from the closed intestinal loops of both jejunal and ileal regions was analyzed for clioquinol and the metabolites and it was found that clioquinol glucuronide was formed predominantly in both regions. From the results of the present studies, intestinal metabolism of clioquinol can be pointed out as a major factor for difficulty to cause clioquinol intoxication.

The existence of the enterohepatic circulation (EHC) of clioquinol was confirmed by using paired rats, donor and recipient, which were connected to each other with a bile duct-to-duodenum cannula. The concentrations of clioquinol and its metabolites appearing in the plasma of the recipient following intraduodenal 10 mg/kg dose of clioquinol to the donor were fairly low. However, within 24 h after the administration ca. 12% of the dose was reexcreted in the bile of the recipient as clioquinol glucuronide and ca. 2% in the urine as clioquinol sulfate. From these results and the data of biliary excretion in our previous paper, the glucuronide was found to play a role on the EHC. Further, both in vitro and in situ results suggested that clioquinol glucuronide excreted in the bile may be absorbed partially after return to the parent drug in the intestinal tract and partially as such without deconjugation.

In order to establish the role of human papillomavirus (HPV) in carcinogenesis of epidermodysplasia verruciformis (EV), the presence, the molecular characteristics and the physical state of HPV DNA in a benign lesion, a primary carcinoma and a metastatic carcinoma developing in the same EV patient were studied and compared. Of the 2 HPV DNAs isolated from benign macular lesions, only one (a subtype of HPV 5) was detected in both primary and metastatic tumors. Only one normal species of viral DNA molecule was detected in the benign lesion, whereas most, if not all, viral DNA molecules present in the carcinoma (both primary and metastatic) were aberrant ones. The major viral DNA molecule in the primary carcinoma was a large HPV DNA with duplicated 40% subgenomic segments, and was present as free episomes. The major viral DNA molecule in the metastatic carcinoma was the 40% subgenomic segment itself, lacking the remaining 60% segment of the viral genome, and was integrated within cellular DNA. Thus, HPV DNA was present in tumors at any stage of malignancy, and its molecular characteristics and physical state changed not only with the development but also with the enhancement of malignancy, consistently conserving its defined 40% subgenomic segment as the predominant viral sequences. Our results suggest that HPV 5 may be actually involved in carcinogenesis in EV patients.

The investigation was undertaken to study the neurological symptoms in rats caused by maintaining high plasma concentration of about 30 nmol/ml or more, of clioquinol. Clioquinol suspension which was prepared using polysorbate 80 was administered intraperitoneally to rats and plasma and tissue concentrations were determined. On administration of clioquinol of 100 and 200 mg/kg, the mean plasma concentrations of clioquinol reached maximum values of 30 and 58 nmol/ml, respectively, after 0.5-1 h and thereafter decreased rapidly. With 400 mg/kg, however, plasma concentration reached maximum value of about 75 nmol/ml and fell slowly. By single and repeated administration of the suspension, clioquinol was distributed in the liver and kidney at a high concentration, and also in the nervous system. In experiments on appearance of neurotoxicity in rats by repeated administration of the suspension, all of 10 rats administered intraperitoneally with 100 mg/kg/d did not develop any neurological symptoms for about 30 d. On the other hand, one of 10 and 7 of 13 rats administered with 200 and 400 mg/kg/d, respectively, developed ataxia in the hind legs or all legs on the 3rd to the 12th day after starting administration. Pathologically, a slight change of the peripheral nerve, central chromatolysis of the anterior horn neuron and severe neuronal degeneration of the Ammon's horn were observed in the rats with ataxia.

Ten hemangioblastomas of the central nervous system were examined by an immunoperoxidase technique for glial fibrillary acidic protein (GFAP), S-100 protein, and Factor VIII-related antigen to determine the origin of the stromal cells. In seven cases scattered stromal cells were positive for GFAP, and eight cases showed variable numbers of S-100 protein-positive cells. These positive cells were present throughout the tumors. S-100 protein-positive stromal cells corresponded to GFAP-positive cells, but the former were more numerous than the latter. Staining for Factor VIII-related antigen was negative in stromal cells, except in endothelial cells lining capillaries. It was suggested that stromal cells of capillary hemangioblastomas are composed by heterogeneous cell populations.

Choriocarcinoma was found in the lung of a 34-year-old woman. Examination of the patient's entire body, especially the genital tract, failed to disclose foci of choriocarcinoma other than that in the right lung. After surgery, the levels of human chorionic gonadotropin in the blood and the urine fell. It was concluded that the choriocarcinoma of the lung was, in fact, the primary tumor. The genesis of choriocarcinoma was also studied in ten patients who died after delivery or abortion. Autopsy disclosed trophoblasts in the pulmonary arteries in nine of these ten patients. These findings suggest that primary choriocarcinoma in women is due to pulmonary embolism caused by trophoblasts at the time of abortion or delivery.

Two distinct human papillomavirus (HPV) DNAs (MY-1 and MY-2) were molecularly cloned from the benign skin lesions of a patient with epidermodysplasia verruciformis. The restriction map of MY-1 was the same as that of HPV 3a. The map of MY-2 appeared to be different from those of any HPVs reported in the literature. MY-2 did not cross-hybridize with MY-1 or the DNAs of HPV types 1, 2 and 4 under stringent conditions.