Three ancient documents solve the jigsaw of the parchment purple spot deterioration and validate the microbial succession model.

The preservation of cultural heritage is one of the major challenges of today's society. Parchments, a semi-solid matrix of collagen produced from animal skin, are a significant part of the cultural heritage, being used as writing material since ancient times. Due to their animal origin, parchments easily undergo biodeterioration: the most common biological damage is characterized by isolated or coalescent purple spots, that often lead to the detachment of the superficial layer and the consequent loss of written content. Although many parchments with purple spot biodegradative features were studied, no common causative agent had been identified so far. In a previous study a successional model has been proposed, basing on the multidisciplinary analysis of damaged versus undamaged samples from a moderately damaged document. Although no specific sequences were observed, the results pointed to Halobacterium salinarum as the starting actor of the succession. In this study, to further investigate this topic, three dramatically damaged parchments were analysed; belonging to a collection archived as Faldone Patrizi A 19, and dated back XVI-XVII century A.D. With the same multidisciplinary approach, the Next Generation Sequencing (NGS, Illumina platform) revealed DNA sequences belonging to Halobacterium salinarum; the RAMAN spectroscopy identified the pigment within the purple spots as haloarchaeal bacterioruberin and bacteriorhodopsine, and the LTA technique quantified the extremely damaged collagen structures through the entire parchments, due to the biological attack to the parchment frame structures. These results allowed to propose a model of the progressive degradation pattern of the parchment collagen. Overall, these data validate a multi-phase microbial succession model. This demonstration is pivotal to possible new restoration strategies, important for a huge number of ancient documents.

Radiation resistance in thermophiles: mechanisms and applications.

The study of prokaryotic life in high temperature environments viz., geothermal areas, hot, acidic geysers and undersea hydrothermal vents has revealed the existence of thermophiles (or hyperthermophiles). These microorganisms possess various stress adaptation mechanisms which enable them to bypass multiple physical and chemical barriers for survival. The discovery of radiation resistant thermophile Deinococcus geothermalis has given new insights into the field of radiation microbiology. The ability of radiation resistant thermophiles to deal with the lethal effects of ionizing radiations like DNA damage, oxidative bursts and protein damage has made them a model system for exobiology and interplanetary transmission of life. They might be an antiquity of historical transport process that brought microbial life on Earth. These radiation resistant thermophiles are resistant to desiccation as well and maintain their homeostasis by advance DNA repair mechanisms, reactive oxygen species (ROS) detoxification system and accumulation of compatible solutes. Moreover, engineered radioresistant thermophilic strains are the best candidate for bioremediation of radionuclide waste while the extremolytes produced by these organisms may have predicted therapeutic uses. So, the present article delineate a picture of radiation resistance thermophiles, their adaptive mechanisms to evade stress viz., radiation and desiccation, their present applications along with new horizons in near future.

Halobacterial nano vesicles displaying murine bactericidal permeability-increasing protein rescue mice from lethal endotoxic shock.

Bactericidal/permeability-increasing protein (BPI) had been shown to possess anti-inflammatory and endotoxin neutralizing activity by interacting with LPS of Gram-negative bacteria. The current study examines the feasibility of using murine BPI (mBPI) expressed on halophilic Archaeal gas vesicle nanoparticles (GVNPs) for the treatment of endotoxemia in high-risk patients, using a murine model of D-galactosamine-induced endotoxic shock. Halobacterium sp. NRC-1was used to express the N-terminal 199 amino acid residues of mBPI fused to the GVNP GvpC protein, and bound to the surface of the haloarchaeal GVNPs. Our results indicate that delivery of mBPIN-GVNPs increase the survival rate of mice challenged with lethal concentrations of lipopolysaccharide (LPS) and D-galactosamine. Additionally, the mBPIN-GVNP-treated mice displayed reduced symptoms of inflammation, including inflammatory anemia, recruitment of neutrophils, liver apoptosis as well as increased pro-inflammatory serum cytokine levels.

Genomic Analysis of the Extremely Halophilic Archaeon Halobacterium noricense CBA1132 Isolated from Solar Salt That Is an Essential Material for Fermented Foods.

The extremely halophilic archaeon Halobacterium noricense is a member of the genus Halobacterium. Strain CBA1132 (= KCCM 43183, JCM 31150) was isolated from solar salt. The genome of strain CBA1132 assembled with 4 contigs, including three rRNA genes, 44 tRNA genes, and 3,208 open reading frames. Strain CBA1132 had nine putative CRISPRs and the genome contained genes encoding metal resistance determinants: copper-translocating P-type ATPase (CtpA), arsenical pump-driving ATPase (ArsA), arsenate reductase (ArsC), and arsenical resistance operon repressor (ArsR). Strain CBA1132 was related to Halobacterium noricense, with 99.2% 16S rRNA gene sequence similarity. Based on the comparative genomic analysis, strain CBA1132 has distinctly evolved; moreover, essential genes related to nitrogen metabolism were only detected in the genome of strain CBA1132 among the reported genomes in the genus Halobacterium. This genome sequence of Halobacterium noricense CBA1132 may be of use in future molecular biological studies.

Halobacterium rubrum sp. nov., isolated from a marine solar saltern.

Halophilic archaeal strain TGN-42-S1(T) was isolated from the Tanggu marine solar saltern, China. Cells from strain TGN-42-S1(T) were observed to be pleomorphic rods, stained Gram-negative, and formed red-pigmented colonies on solid media. Strain TGN-42-S1(T) was found to be able to grow at 20-50 °C (optimum 35-37 °C), at 1.7-4.8 M NaCl (optimum 3.1 M), at 0-1.0 M MgCl2 (optimum 0.1 M), and at pH 5.0-9.0 (optimum pH 7.0-7.5). The cells lysed in distilled water, and the minimal NaCl concentration to prevent cell-lysis was found to be 10 % (w/v). The major polar lipids of the strain were phosphatidic acid, phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol sulfate, galactosyl mannosyl glucosyl diether (TGD-1), sulfated galactosyl mannosyl glucosyl diether (S-TGD-1), sulfated galactosyl mannosyl galactofuranosyl glucosyl diether (S-TeGD), and three unidentified glycolipids which were chromatographically identical to those of the Halobacterium species. The 16S rRNA gene and rpoB' gene of strain TGN-42-S1(T) were phylogenetically related to the corresponding genes of Halobacterium jilantaiense CGMCC 1.5337(T) (98.8 and 93.5 % nucleotide identity, respectively), Halobacterium salinarum CGMCC 1.1958(T) (98.4 and 91.9 %), and Halobacterium noricense JCM 15102(T) (96.9 and 91.1 %). The DNA G + C content of strain TGN-42-S1(T) was determined to be 69.2 mol %. Strain TGN-42-S1(T) showed low DNA-DNA relatedness with Hbt. jilantaiense CGMCC 1.5337(T) and Hbt. salinarum CGMCC 1.1958(T), the most closely related members of the genus Halobacterium. The phenotypic, chemotaxonomic, and phylogenetic properties suggested that strain TGN-42-S1(T) (=CGMCC 1.12575(T) =JCM 19908(T)) represents a new species of Halobacterium, for which the name Halobacterium rubrum sp. nov. is proposed.

Hyperhalophilic archaeal biofilms: growth kinetics, structure, and antagonistic interaction in continuous culture.

Biofilms by the hyperhalophilic archaea Halorubrum sp. and Halobacterium sp. were analyzed, and for the first time the progression of structural features and the developmental parameters of these sessile populations are described. Optical slicing and digital analysis of sequential micrographs showed that their three dimensional structure was microorganism dependent. Biofilms of Halobacterium sp. developed in clusters that covered about 30% of the supporting surface at the interface level and expanded over about 86 ± 4 µm in thickness, while Halorubrum sp. biofilms covered less than 20% of the surface and reached a thickness of 41 ± 1 µm. The kinetics of growth was lower in biofilms, with generation times of 27 ± 1 and 36 ± 2 h for Halobacterium sp. and Halorubrum sp., respectively, as compared to 8.4 ± 0.3 and 14 ± 1 h in planktonic cultures. Differences between microorganisms were also observed at the cell morphology level. The interaction between the two microorganisms was also evaluated, showing that Halobacterium sp. can outcompete already established Halorubrum sp. biofilms by a mechanism that might include the combined action of tunnelling swimmers and antimicrobial compounds.

The protein interaction network of a taxis signal transduction system in a halophilic archaeon.

BACKGROUND: The taxis signaling system of the extreme halophilic archaeon Halobacterium (Hbt.) salinarum differs in several aspects from its model bacterial counterparts Escherichia coli and Bacillus subtilis. We studied the protein interactions in the Hbt. salinarum taxis signaling system to gain an understanding of its structure, to gain knowledge about its known components and to search for new members. RESULTS: The interaction analysis revealed that the core signaling proteins are involved in different protein complexes and our data provide evidence for dynamic interchanges between them. Fifteen of the eighteen taxis receptors (halobacterial transducers, Htrs) can be assigned to four different groups depending on their interactions with the core signaling proteins. Only one of these groups, which contains six of the eight Htrs with known signals, shows the composition expected for signaling complexes (receptor, kinase CheA, adaptor CheW, response regulator CheY). From the two Hbt. salinarum CheW proteins, only CheW1 is engaged in signaling complexes with Htrs and CheA, whereas CheW2 interacts with Htrs but not with CheA. CheY connects the core signaling structure to a subnetwork consisting of the two CheF proteins (which build a link to the flagellar apparatus), CheD (the hub of the subnetwork), two CheC complexes and the receptor methylesterase CheB. CONCLUSIONS: Based on our findings, we propose two hypotheses. First, Hbt. salinarum might have the capability to dynamically adjust the impact of certain Htrs or Htr clusters depending on its current needs or environmental conditions. Secondly, we propose a hypothetical feedback loop from the response regulator to Htr methylation made from the CheC proteins, CheD and CheB, which might contribute to adaptation analogous to the CheC/CheD system of B. subtilis.

Genome-wide responses of the model archaeon Halobacterium sp. strain NRC-1 to oxygen limitation.

As part of a comprehensive postgenomic investigation of the model archaeon Halobacterium sp. strain NRC-1, we used whole-genome DNA microarrays to compare transcriptional profiles of cells grown under anaerobic or aerobic conditions. When anaerobic growth supported by arginine fermentation was compared to aerobic growth, genes for arginine fermentation (arc) and anaerobic respiration (dms), using trimethylamine N-oxide (TMAO) as the terminal electron acceptor, were highly upregulated, as was the bop gene, required for phototrophic growth. When arginine fermentation was compared to anaerobic respiration with TMAO, the arc and dms genes were both induced with arginine, while TMAO induced the bop gene and major gas vesicle protein (gvpAC) genes specifying buoyant gas vesicles. Anaerobic conditions with either TMAO or arginine also upregulated the cba genes, encoding one of three cytochrome oxidases. In-frame deletion of two COG3413 family regulatory genes, bat and dmsR, showed downregulation of the bop gene cluster and loss of purple membrane synthesis and downregulation of the dms operon and loss of anaerobic respiration capability, respectively. Bioinformatic analysis identified additional regulatory and sensor genes that are likely involved in the full range of cellular responses to oxygen limitation. Our results show that the Halobacterium sp. has evolved a carefully orchestrated set of responses to oxygen limitation. As conditions become more reducing, cells progressively increase buoyancy, as well as capabilities for phototrophy, scavenging of molecular oxygen, anaerobic respiration, and fermentation.

The Inferelator 2.0: a scalable framework for reconstruction of dynamic regulatory network models.

Current methods for reconstructing biological networks often learn either the topology of large networks or the kinetic parameters of smaller networks with a well-characterized topology. We have recently described a network reconstruction algorithm, the Inferelator 1.0, that given a set of genome-wide measurements as input, simultaneously learns both topology and kinetic-parameters. Specifically, it learns a system of ordinary differential equations (ODEs) that describe the rate of change in transcription of each gene or gene-cluster, as a function of environmental and transcription factors. In order to scale to large networks, in Inferelator 1.0 we have approximated the system of ODEs to be uncoupled, and have solved each ODE using a one-step finite difference approximation. Naturally, these approximations become crude as the simulated time-interval increases. Here we present, implement, and test a new Markov-Chain-Monte-Carlo (MCMC) dynamical modeling method, Inferelator 2.0, that works in tandem with Inferelator 1.0 and is designed to relax these approximations. We show results for the prokaryote Halobacterium that demonstrate a marked improvement in our predictive performance in modeling the regulatory dynamics of the system over longer time-scales.

Quantitative analysis of signal transduction in motile and phototactic cells by computerized light stimulation and model based tracking.

To investigate the responses of Halobacterium salinarum to stimulation with light (phototaxis and photokinesis), we designed an experimental setup consisting of optical devices for automatic video image acquisition and computer-controlled light stimulation, and developed algorithms to analyze physiological responses of the cells. Cells are categorized as motile and nonmotile by a classification scheme based on the square displacement of cell positions. Computerized tracking based on a dynamic model of the stochastic cell movement and a Kalman filter-based algorithm allows smoothed estimates of the cell tracks and the detection of physiological responses to complex stimulus patterns. The setup and algorithms were calibrated which allows quantitative measurements and systematic analysis of cellular sensing and response. Overall, the setup is flexible, extensible, and consists mainly of commercially available products. This facilitates modifications of the setup and algorithms for physiological studies of the motility of cells or microorganisms.

Halobacterium piscisalsi sp. nov., from fermented fish (pla-ra) in Thailand.

A Gram-negative, motile, rod-shaped, extremely halophilic archaeon, designated strain HPC1-2(T), was isolated from pla-ra, a salt-fermented fish product of Thailand. Strain HPC1-2(T) was able to grow at 20-60 degrees C (optimum at 37-40 degrees C), at 2.6-5.1 M NaCl (optimum at 3.4-4.3 M NaCl) and at pH 5.0-8.0 (optimum at pH 7.0-7.5). Hypotonic treatment with less than 1.7 M NaCl caused cell lysis. The major polar lipids of the isolate were C(20)C(20) derivatives of phosphatidylglycerol, methylated phosphatidylglycerol phosphate, phosphatidylglycerol sulfate, triglycosyl diether, sulfated triglycosyl diether and sulfated tetraglycosyl diether. The G+C content of the DNA was 65.5 mol%. 16S rRNA gene sequence analysis indicated that the isolate represented a member of the genus Halobacterium in the family Halobacteriaceae. Based on 16S rRNA gene sequence similarity, strain HPC1-2(T) was related most closely to Halobacterium salinarum DSM 3754(T) (99.2%) and Halobacterium jilantaiense JCM 13558(T) (97.8%). However, low levels of DNA-DNA relatedness suggested that strain HPC1-2(T) was genotypically different from these closely related type strains. Strain HPC1-2(T) could also be differentiated based on physiological and biochemical characteristics. Therefore, strain HPC1-2(T) is considered to represent a novel species of the genus Halobacterium, for which the name Halobacterium piscisalsi sp. nov. is proposed. The type strain is HPC1-2(T) (=BCC 24372(T)=JCM 14661(T)=PCU 302(T)).

Rad50 is not essential for the Mre11-dependent repair of DNA double-strand breaks in Halobacterium sp. strain NRC-1.

The genome of the halophilic archaeon Halobacterium sp. strain NRC-1 encodes homologs of the eukaryotic Mre11 and Rad50 proteins, which are involved in the recognition and end processing of DNA double-strand breaks in the homologous recombination repair pathway. We have analyzed the phenotype of Halobacterium deletion mutants lacking mre11 and/or rad50 after exposure to UV-C radiation, an alkylating agent (N-methyl-N'-nitro-N-nitrosoguanidine), and gamma radiation, none of which resulted in a decrease in survival of the mutant strains compared to that of the background strain. However, a decreased rate of repair of DNA double-strand breaks in strains lacking the mre11 gene was observed using pulsed-field gel electrophoresis. These observations led to the hypothesis that Mre11 is essential for the repair of DNA double-strand breaks in Halobacterium, whereas Rad50 is dispensable. This is the first identification of a Rad50-independent function for the Mre11 protein, and it represents a shift in the Archaea away from the eukaryotic model of homologous recombination repair of DNA double-strand breaks.

An integrated systems approach for understanding cellular responses to gamma radiation.

Cellular response to stress entails complex mRNA and protein abundance changes, which translate into physiological adjustments to maintain homeostasis as well as to repair and minimize damage to cellular components. We have characterized the response of the halophilic archaeon Halobacterium salinarum NRC-1 to (60)Co ionizing gamma radiation in an effort to understand the correlation between genetic information processing and physiological change. The physiological response model we have constructed is based on integrated analysis of temporal changes in global mRNA and protein abundance along with protein-DNA interactions and evolutionarily conserved functional associations. This systems view reveals cooperation among several cellular processes including DNA repair, increased protein turnover, apparent shifts in metabolism to favor nucleotide biosynthesis and an overall effort to repair oxidative damage. Further, we demonstrate the importance of time dimension while correlating mRNA and protein levels and suggest that steady-state comparisons may be misleading while assessing dynamics of genetic information processing across transcription and translation.

A systems view of haloarchaeal strategies to withstand stress from transition metals.

Given that transition metals are essential cofactors in central biological processes, misallocation of the wrong metal ion to a metalloprotein can have resounding and often detrimental effects on diverse aspects of cellular physiology. Therefore, in an attempt to characterize unique and shared responses to chemically similar metals, we have reconstructed physiological behaviors of Halobacterium NRC-1, an archaeal halophile, in sublethal levels of Mn(II), Fe(II), Co(II), Ni(II), Cu(II), and Zn(II). Over 20% of all genes responded transiently within minutes of exposure to Fe(II), perhaps reflecting immediate large-scale physiological adjustments to maintain homeostasis. At steady state, each transition metal induced growth arrest, attempts to minimize oxidative stress, toxic ion scavenging, increased protein turnover and DNA repair, and modulation of active ion transport. While several of these constitute generalized stress responses, up-regulation of active efflux of Co(II), Ni(II), Cu(II), and Zn(II), down-regulation of Mn(II) uptake and up-regulation of Fe(II) chelation, confer resistance to the respective metals. We have synthesized all of these discoveries into a unified systems-level model to provide an integrated perspective of responses to six transition metals with emphasis on experimentally verified regulatory mechanisms. Finally, through comparisons across global transcriptional responses to different metals, we provide insights into putative in vivo metal selectivity of metalloregulatory proteins and demonstrate that a systems approach can help rapidly unravel novel metabolic potential and regulatory programs of poorly studied organisms.

Gas vesicles isolated from Halobacterium cells by lysis in hypotonic solution are structurally weakened.

Analysis of pressure-collapse curves of Halobacterium cells containing gas vesicles and of gas vesicles released from such cells by hypotonic lysis shows that the isolated gas vesicles are considerably weaker than those present within the cells: their mean critical collapse pressure was around 0.049-0.058 MPa, as compared to 0.082-0.095 MPa for intact cells. The hypotonic lysis procedure, which is widely used for the isolation of gas vesicles from members of the Halobacteriaceae, thus damages the mechanical properties of the vesicles. The phenomenon can possibly be attributed to the loss of one or more structural gas vesicle proteins such as GvpC, the protein that strengthens the vesicles built of GvpA subunits: Halobacterium GvpC is a highly acidic, typically "halophilic" protein, expected to denature in the absence of molar concentrations of salt.

A quantitative model of the switch cycle of an archaeal flagellar motor and its sensory control.

By reverse-engineering we have detected eight kinetic phases of the symmetric switch cycle of the Halobacterium salinarum flagellar motor assembly and identified those steps in the switch cycle that are controlled by sensory rhodopsins during phototaxis. Upon switching the rotational sense, the flagellar motor assembly passes through a stop state from which all subunits synchronously resume rotation in the reverse direction. The assembly then synchronously proceeds through three subsequent functional states of the switch: Refractory, Competent, and Active, from which the rotational sense is switched again. Sensory control of the symmetric switch cycle occurs at two steps in each rotational sense by inversely regulating the probabilities for a change from the Refractory to the Competent and from Competent to the Active rotational mode. We provide a mathematical model for flagellar motor switching and its sensory control, which is able to explain all tested experimental results on spontaneous and light-controlled motor switching, and give a mechanistic explanation based on synchronous conformational transitions of the subunits of the switch complex after reversible dissociation and binding of a response regulator (CheYP). We conclude that the kinetic mechanism of flagellar motor switching and its sensory control is fundamentally different in the archaeon H. salinarum and the bacterium Escherichia coli.

Physiological responses of the halophilic archaeon Halobacterium sp. strain NRC1 to desiccation and gamma irradiation.

We report that the halophilic archaeon Halobacterium sp. strain NRC-1 is highly resistant to desiccation, high vacuum and 60Co gamma irradiation. Halobacterium sp. was able to repair extensive double strand DNA breaks (DSBs) in its genomic DNA, produced both by desiccation and by gamma irradiation, within hours of damage induction. We propose that resistance to high vacuum and 60Co gamma irradiation is a consequence of its adaptation to desiccating conditions. Gamma resistance in Halobacterium sp. was dependent on growth stage with cultures in earlier stages exhibiting higher resistance. Membrane pigments, specifically bacterioruberin, offered protection against cellular damages induced by high doses (5 kGy) of gamma irradiation. High-salt conditions were found to create a protective environment against gamma irradiation in vivo by comparing the amount of DSBs induced by ionizing radiation in the chromosomal DNA of Halobacterium sp. to that of the more radiation-sensitive Escherichia coli that grows in lower-salt conditions. No inducible response was observed after exposing Halobacterium sp. to a nonlethal dose (0.5 kGy) of gamma ray and subsequently exposing the cells to either a high dose (5 kGy) of gamma ray or desiccating conditions. We find that the hypersaline environment in which Halobacterium sp. flourishes is a fundamental factor for its resistance to desiccation, damaging radiation and high vacuum.

Cassette-based presentation of SIV epitopes with recombinant gas vesicles from halophilic archaea.

In earlier studies we demonstrated recombinant gas vesicles from Halobacterium sp. NRC-1, expressing a model six amino acid insert, or native vesicles displaying chemically coupled TNP, each were immunogenic, and antigenic. Long-lived responses displaying immunologic memory were elicited without exogenous adjuvant. Here we report the generation and expression of cassettes containing SIV derived DNA. The results indicate a cassette-based display/delivery system derived from recombinant halobacterial gas vesicle genes is highly feasible. Data specifically support four conclusions: (i) Recombinants carrying up to 705 bp of SIV DNA inserted into the gvpC gene form functional gas vesicles; (ii) SIV peptides contained as part of the expressed recombinant, surface exposed GvpC protein are recognized by antibody elicited in monkeys exposed to native SIV in vivo; (iii) in the absence of adjuvant, mice immunized with the recombinant gas vesicle (r-GV) preparations mount a solid, titratable antibody response to the test SIV insert that is long lived and exhibits immunologic memory; (iv) recombinant organelles, created through the generation of cassettes encoding epitopes inserted into the gvpC DNA, can be used to construct a multiepitope display (MED) library, a potentially cost effective vehicle to express and deliver peptides of SIV, HIV or other pathogens.

Cytoscape: a software environment for integrated models of biomolecular interaction networks.

Cytoscape is an open source software project for integrating biomolecular interaction networks with high-throughput expression data and other molecular states into a unified conceptual framework. Although applicable to any system of molecular components and interactions, Cytoscape is most powerful when used in conjunction with large databases of protein-protein, protein-DNA, and genetic interactions that are increasingly available for humans and model organisms. Cytoscape's software Core provides basic functionality to layout and query the network; to visually integrate the network with expression profiles, phenotypes, and other molecular states; and to link the network to databases of functional annotations. The Core is extensible through a straightforward plug-in architecture, allowing rapid development of additional computational analyses and features. Several case studies of Cytoscape plug-ins are surveyed, including a search for interaction pathways correlating with changes in gene expression, a study of protein complexes involved in cellular recovery to DNA damage, inference of a combined physical/functional interaction network for Halobacterium, and an interface to detailed stochastic/kinetic gene regulatory models.

FTIR spectroscopy of the complex between pharaonis phoborhodopsin and its transducer protein.

pharaonis phoborhodopsin (ppR; also called pharaonis sensory rhodopsin II, psRII) is a photoreceptor for negative phototaxis in Natronobacterium pharaonis. ppR activates the cognate transducer protein, pHtrII, upon absorption of light. ppR and pHtrII form a tight 2:2 complex in the unphotolyzed state, and the interaction is somehow altered during the photocycle of ppR. In this paper, we studied the influence of pHtrII on the structural changes occurring upon retinal photoisomerization in ppR by means of low-temperature FTIR spectroscopy. We trapped the K intermediate at 77 K and compared the ppR(K) minus ppR spectra in the absence and presence of pHtrII. There are no differences in the X-D stretching vibrations (2700-1900 cm(-1)) caused by presence of pHtrII. This result indicates that the hydrogen-bonding network in the Schiff base region is not altered by interaction with pHtrII, which is consistent with the same absorption spectrum of ppR with or without pHtrII. In contrast, the ppR(K) minus ppR infrared difference spectra are clearly influenced by the presence of pHtrII in amide-I (1680-1640 cm(-1)) and amide-A (3350-3250 cm(-1)) vibrations. The identical spectra for the complex of the unlabeled ppR and (13)C- or (15)N-labeled pHtrII indicate that the observed structural changes for the peptide backbone originate from ppR only and are altered by retinal photoisomerization. The changes do not come from pHtrII, implying that the light signal is not transmitted to pHtrII in ppR(K). In addition, we observed D(2)O-insensitive bands at 3479 (-)/3369 (+) cm(-1) only in the presence of pHtrII, which presumably originate from an X-H stretch of an amino acid side chain inside the protein.