Discovery of Substituted 5-(2-Hydroxybenzoyl)-2-Pyridone Analogues as Inhibitors of the Human Caf1/CNOT7 Ribonuclease.

The Caf1/CNOT7 nuclease is a catalytic component of the Ccr4-Not deadenylase complex, which is a key regulator of post-transcriptional gene regulation. In addition to providing catalytic activity, Caf1/CNOT7 and its paralogue Caf1/CNOT8 also contribute a structural function by mediating interactions between the large, non-catalytic subunit CNOT1, which forms the backbone of the Ccr4-Not complex and the second nuclease subunit Ccr4 (CNOT6/CNOT6L). To facilitate investigations into the role of Caf1/CNOT7 in gene regulation, we aimed to discover and develop non-nucleoside inhibitors of the enzyme. Here, we disclose that the tri-substituted 2-pyridone compound 5-(5-bromo-2-hydroxy-benzoyl)-1-(4-chloro-2-methoxy-5-methyl-phenyl)-2-oxo-pyridine-3-carbonitrile is an inhibitor of the Caf1/CNOT7 nuclease. Using a fluorescence-based nuclease assay, the activity of 16 structural analogues was determined, which predominantly explored substituents on the 1-phenyl group. While no compound with higher potency was identified among this set of structural analogues, the lowest potency was observed with the analogue lacking substituents on the 1-phenyl group. This indicates that substituents on the 1-phenyl group contribute significantly to binding. To identify possible binding modes of the inhibitors, molecular docking was carried out. This analysis suggested that the binding modes of the five most potent inhibitors may display similar conformations upon binding active site residues. Possible interactions include π-π interactions with His225, hydrogen bonding with the backbone of Phe43 and Van der Waals interactions with His225, Leu209, Leu112 and Leu115.

A fluorescence-based assay suitable for quantitative analysis of deadenylase enzyme activity.

In eukaryotic cells, the shortening and removal of the poly(A) tail of cytoplasmic mRNA by deadenylase enzymes is a critical step in post-transcriptional gene regulation. The ribonuclease activity of deadenylase enzymes is attributed to either a DEDD (Asp-Glu-Asp-Asp) or an endonuclease-exonuclease-phosphatase domain. Both domains require the presence of two Mg2+ ions in the active site. To facilitate the biochemical analysis of deadenylase enzymes, we have developed a fluorescence-based deadenylase assay. The assay is based on end-point measurement, suitable for quantitative analysis and can be adapted for 96- and 384-well microplate formats. We demonstrate the utility of the assay by screening a chemical compound library, resulting in the identification of non-nucleoside inhibitors of the Caf1/CNOT7 enzyme, a catalytic subunit of the Ccr4-Not deadenylase complex. These compounds may be useful tools for the biochemical analysis of the Caf1/CNOT7 deadenylase subunit of the Ccr4-Not complex and indicate the feasibility of developing selective inhibitors of deadenylase enzymes using the fluorescence-based assay.

Discovery, synthesis and biochemical profiling of purine-2,6-dione derivatives as inhibitors of the human poly(A)-selective ribonuclease Caf1.

Eukaryotic mRNA contains a 3' poly(A) tail, which plays important roles in the regulation of mRNA stability and translation. Well-characterized enzymes involved in the shortening of the poly(A) tail include the multi-subunit Ccr4-Not deadenylase, which contains the Caf1 (Pop2) and Ccr4 catalytic components, and poly(A)-specific ribonuclease (PARN). Two Mg(2+) ions present in the active sites of these ribonucleases are required for RNA cleavage. Here, we report the discovery, synthesis and biochemical profiling of purine-2,6-dione derivatives as (sub)micromolar inhibitors of Caf1.

Exiguobacterium himgiriensis sp. nov. a novel member of the genus Exiguobacterium, isolated from the Indian Himalayas.

The taxonomic position of an orange coloured bacterium, strain K22-26(T) isolated from a soil sample was studied using a polyphasic approach. The organism had phenotypic and chemotaxonomic properties consistent with its allocation into the genus Exiguobacterium. Phylogenetic analysis of the 16S rRNA gene sequence showed that strain K22-26(T) belongs to the genus Exiguobacterium and was related to Exiguobacterium aurantiacum DSM 6208(T) (99.0 %) Exiguobacterium mexicanum DSM 16483(T) (98.6 %), Exiguobacterium aquaticum (98.6 %), Exiguobacterium aestuarii DSM 16306(T) (98.1 %), Exiguobacterium profundum DSM 17289(T) (98.1 %) and Exiguobacterium marinum DSM 16483(T) (97.9 %), whereas sequence similarity values with respect to other Exiguobacterium species with validly published names were between 92.5-94.0 %. The major polar lipids detected were phosphatidylglycerol, diphosphatidylglycerol and phosphatidylethanolamine. The major menaquinone was determined to be MK-7 (83 %) whereas MK-8 (11 %) and MK-6 (6 %) occur in smaller amounts. The peptidoglycan of the strain was found to contain L-lysine as the diagnostic diamino acid. The major fatty acids detected were iso C13:0 (11.2 %), anteiso C13:0 (15.4 %), iso C15:0 (13.2 %) and iso C17:0 (16.1 %). However, analysis of the DNA-DNA relatedness confirmed that strain K22-26(T) belongs to a novel species. The G + C content of the strain K22-26(T) was determined to be 50.1 mol %. The novel strain was distinguished from closely related type species of the genus Exiguobacterium using DNA-DNA relatedness and phenotypic data. Based on these differences, the strain K22-26(T) should be classified as a novel species of the genus Exiguobacterium, for which the name Exiguobacterium himgiriensis sp. nov. strain K22-26(T) (= MTCC 7628(T) = JCM 14260(T)) is proposed.

Flavobacterium rakeshii sp. nov., isolated from marine sediment, and emended description of Flavobacterium beibuense Fu et al. 2011.

A Gram-negative, non-motile bacterial strain that formed straight rods and straw yellow colonies, designated FCS-5(T), was isolated from a marine sediment from the Arabian Sea. The isolate exhibited most of the phenotypic properties expected for a member of the genus Flavobacterium. The major fatty acids were iso-C(15:0), iso-C(17:0) 3-OH, C(17:1)ω9c and summed feature 3 (comprising iso-C(15:0) 2-OH and/or C(16:1)ω7c). The only isoprenoid quinone was MK-6. The only polyamine was homospermidine and the major polar lipid was phosphatidylethanolamine. The G+C content of the genomic DNA was 32.4 mol%. According to 16S rRNA gene sequence analysis, strain FCS-5(T) belonged to the genus Flavobacterium and exhibited 99.3% 16S rRNA gene sequence similarity with Flavobacterium beibuense F44-8(T) and 90.9-94.6% sequence similarity with other members of the genus Flavobacterium. The results of physiological and biochemical tests allowed the discrimination of the isolate from its phylogenetic relatives. Strain FCS-5(T) is a representative of a novel species of the genus Flavobacterium, for which the name Flavobacterium rakeshii sp. nov. is proposed. The type strain is FCS-5(T) ( = MTCC 10967(T) = JCM 17928(T)). An emended description of F. beibuense is also proposed.

Planococcus plakortidis sp. nov., isolated from the marine sponge Plakortis simplex (Schulze).

A novel coccoid-shaped strain, AS/ASP6 (II)T, was isolated from a sample taken from Plakortis simplex (Schulze), a marine sponge, collected at a depth of 30 m from the Bay of Bengal. This strain was identified by using a polyphasic taxonomic approach. The results of 16S rRNA gene sequence analysis showed that strain AS/ASP6 (II)T should be assigned to the genus Planococcus. Chemotaxonomic data (A4α-type peptidoglycan; MK-6, MK-7 and MK-8 menaquinones; mainly branched cellular fatty acids; and phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol as cellular phospholipids) supported taxonomic placement in the genus Planococcus. Phylogenetic analysis of the 16S rRNA gene sequence showed that strain AS/ASP6 (II)T belonged to the genus Planococcus and was closely related to the type strains of Planococcus maritimus (99.1 %) followed by Planococcus rifietoensis (98.6 %), Planococcus maitriensis (98.5 %), Planococcus citreus (98.3 %), Planococcus salinarum (98.1 %), Planococcus columbae (97.9 %), Planococcus donghaensis (97.8 %) and Planococcus antarcticus (97.7 %); DNA-DNA hybridization values obtained were well below the threshold that is required for the proposal of a novel species. The G+C content of the genomic DNA was 51.0  mol%. The phenotypic and genotypic data showed that strain AS/ASP6 (II)T merits recognition as a representative of a novel species of the genus Planococcus, for which the name Planococcus plakortidis sp. nov. is proposed; the type strain is AS/ASP6 (II)T (=MTCC 8491T=DSM 23997T).

Nitratireductor lucknowense sp. nov., a novel bacterium isolated from a pesticide contaminated soil.

A straw-yellow pigmented bacterium, strain IITR-21(T) was isolated from a pesticide contaminated site and characterized by using a polyphasic taxonomic approach. The organism had morphological and chemotaxonomic properties consistent with its classification in the genus Nitratireductor. Phylogenetic analysis of the 16S rRNA gene sequence showed that the strain IITR-21(T) belongs to the genus Nitratireductor and was moderately related to Nitratireductor indicus C115(T) (97.7%) and Nitratireductor pacificus pht-3B(T) (97.4%), whereas sequence similarity value with the other species including the type species of the genus Nitratireductor, Nitratireductor aquibiodomus showed less than 97.0% similarity. However, the DNA-DNA relatedness values between strain IITR-21(T) and the moderately related taxa N. indicus (59.1%) and N. pacificus (40.4%) were well below the 70% threshold value recommended for the delineation of bacterial species. The G+C content of the DNA was 62.4 mol%. Based on physiological, biochemical tests and genotypic differences between the strain IITR-21(T) and the other two validly published species of the genus Nitratireductor, it is proposed that the strain be classified as a new species of Nitratireductor as Nitratireductor lucknowense sp. nov. The type strain is IITR-21(T) (=MTCC 8354(T )= DSM 24322(T)).

Kocuria sediminis sp. nov., isolated from a marine sediment sample.

A Gram-positive, pinkish-orange pigmented, coccoid strain, FCS-11(T) was isolated from a marine sediment sample taken from Kochi fort area, Kerala, India and subjected to polyphasic taxonomic study. The 16S rRNA gene sequence of the strain was determined and the results of 16S rRNA gene sequence analysis showed that the strain FCS-11(T) should be assigned to the genus Kocuria. The chemotaxonomic data supported this taxonomic placement i.e. menaquinones MK-7(H(2)), MK-8(H(2)) and MK-9(H(2)); major fatty acids anteiso C15:0 and iso-C15:0 and phosphatidylglycerol (PG) and diphosphatidylglycerol (DPG) as major polar lipids. Further phylogenetic analysis of the 16S rRNA gene sequence confirmed that the strain FCS-11(T) belonged to the genus Kocuria and is closely related to Kocuria turfanensis MTCC 10790(T) (99.4%) followed by Kocuria polaris MTCC 3702(T) (98.2%), Kocuria rosea MTCC 2522(T) (98.2%), Kocuria flava MTCC 10971(T) (98.2%), Kocuria aegyptia MTCC 10791(T) (98.0%), Kocuria himachalensis MTCC 7020(T) (97.5%) and Kocuria atrinae MTCC 10972(T) (97.1%). However, the DNA-DNA hybridisation values obtained between strain FCS-11(T) and other related strains were well below the threshold that is required for the proposal of a novel species. The G+C content of the genomic DNA was 60.7 mol%. The phenotypic and genotypic data showed that the strain FCS-11(T) merits the recognition as a representative of a novel species of the genus Kocuria. It is proposed that the isolate should be classified in the genus Kocuria as a novel species, Kocuria sediminis sp. nov. The type strain is FCS-11(T) (= MTCC 10969(T) = JCM 17929(T)).

Biosynthesis of natural aroma compounds using recombinant whole-cell tomato hydroperoxide lyase biocatalyst.

Green leaf volatiles impart characteristic aroma and flavour to a variety of natural foods due to their inherent grassy note contributed by aldehydes. Hydroperoxide lyase (HPL) is an enzyme that helps in the cleavage of fatty acid hydroperoxides to short-chain aldehydes and ω-oxo-acids. A tomato hydroperoxide lyase gene was successfully expressed in E. coli BL21 (DE3) cells and used in the subsequent production of (Z)-3-hexenal. Biochemical characterization of the HPL activity exhibited by these whole cells enabled the development of a suitable one-pot reaction process for conversion of the hydroperoxide substrate to the corresponding aldehyde, (Z)-3-hexenal, and finally to (Z)-3-hexenol, a high-value flavour and fragrance ingredient.

Interleukin-4-inducing principle from Schistosoma mansoni eggs contains a functional C-terminal nuclear localization signal necessary for nuclear translocation in mammalian cells but not for its uptake.

Interleukin-4-inducing principle from schistosome eggs (IPSE/alpha-1) is a protein produced exclusively by the eggs of the trematode Schistosoma mansoni. IPSE/alpha-1 is a secretory glycoprotein which activates human basophils via an IgE-dependent but non-antigen-specific mechanism. Sequence analyses revealed a potential nuclear localization signal (NLS) at the C terminus of IPSE/alpha-1. Here we show that this sequence (125-PKRRRTY-131) is both necessary and sufficient for nuclear localization of IPSE or IPSE-enhanced green fluorescent protein (EGFP) fusions. While transiently expressed EGFP-IPSE/alpha-1 was exclusively nuclear in the Huh7 and U-2 OS cell lines, a mutant lacking amino acids 125 to 134 showed both nuclear and cytoplasmic staining. Moreover, insertion of the IPSE/alpha-1 NLS into a tetra-EGFP construct rendered the protein nuclear. Alanine scanning mutagenesis revealed a requirement for the KRRR residues. Fluorescence microscopy depicted, and Western blotting further confirmed, that recombinant IPSE/alpha-1 protein added exogenously is rapidly internalized by CHO cells and accumulates in nuclei in an NLS-dependent manner. A mutant protein in which the NLS motif was disrupted by triple mutation (RRR to AAA) was able to penetrate CHO cells but did not translocate to the nucleus. Furthermore, the uptake of native glycosylated IPSE/alpha-1 was confirmed in human primary monocyte-derived dendritic cells and was found to be a calcium- and temperature-dependent process. Live-cell imaging showed that IPSE/alpha-1 is not targeted to lysosomes. In contrast, peripheral blood basophils do not take up IPSE/alpha-1 and do not require the presence of an intact NLS for activation. Taken together, our results suggest that IPSE/alpha-1 may have additional nuclear functions in host cells.

Calidifontibacter indicus gen. nov., sp. nov., a member of the family Dermacoccaceae isolated from a hot spring, and emended description of the family Dermacoccaceae.

During the course of a study on the bacterial diversity in Western Ghats, India, an actinobacterial strain, designated PC IW02(T), was isolated and characterized by a polyphasic taxonomic approach. Strain PC IW02(T) was a non-motile, Gram-positive, short rod that formed creamish white to yellow coloured colonies. 16S rRNA gene sequence analysis showed that the novel strain showed highest sequence similarity with type strains of members of the genus Dermacoccus: Dermacoccus barathri (96.6 %), Dermacoccus profundi (96.5 %), Dermacoccus abyssi (96.4 %) and Dermacoccus nishinomiyaensis (95.9 %). The phylogenetic tree suggested that strain PC IW02(T) could represent a member of a new genus of the family Dermacoccaceae with the genus Demetria as closest clade. Pairwise sequence alignment with Demetria terragena HKI 0089(T) and Kytococcus sedentarius DSM 20547(T) showed similarities of 94.2 and 93.7 %, respectively. Strain PC IW02(T) had MK-8(H(4)) as the major menaquinone. The major fatty acids were iso-C(16 : 0) (43.4 %), iso-C(16 : 1) H (17.2 %) and anteiso-C(17 : 0) (9.9 %). The diagnostic cell-wall amino acid at position 3 of the peptide subunit was lysine; the interpeptide bridge consisted of Gly-Ser-Asp. The polar lipids present were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides and phosphatidylserine, along with two unknown phospholipids. The genomic DNA G+C content of the isolate was 77 mol%. On the basis of phenotypic characteristics, including chemotaxonomic data, and 16S rRNA gene sequence similarities, strain PC IW02(T) represents a novel species in a new genus of the family Dermacoccaceae for which the name Calidifontibacter indicus gen. nov., sp. nov. is proposed. The type strain of Calidifontibacter indicus is PC IW02(T) ( = MTCC 8338(T) = DSM 22967(T) = JCM 16038(T)). An emended description of the family Dermacoccaceae is provided.

Agrococcus carbonis sp. nov., isolated from soil of a coal mine.

An actinobacterial strain, designated G4(T), isolated from a coal mine was subjected to polyphasic taxonomic characterization. Cells were Gram-stain-positive, yellow-pigmented, non-motile and non-spore-forming cocci. This organism possessed a type B peptidoglycan with diaminobutyric acid as diagnostic diamino acid. The major respiratory quinones were MK-9, MK-10 and MK-11. The major fatty acids were anteiso-C(15 : 0) (41.6 %) and anteiso-C(17 : 0) (32.8 %). The predominant cellular polar lipids were diphosphatidylglycerol and phosphatidylglycerol. Cell wall sugars comprised galactose, glucose, ribose and rhamnose. 16S rRNA gene sequence analysis of strain G4(T) showed high similarity with Agrococcus baldri (98.9 %), Agrococcus citreus (97.8 %), Agrococcus jenensis (97.3 %) and Agrococcus terreus (97.0 %). Sequence similarity with the type strains of the other species of the genus Agrococcus was less than 97.0 %. The DNA-DNA relatedness of strain G4(T) with the type strains of Agrococcus baldri, Agrococcus citreus, Agrococcus jenensis and Agrococcus terreus was less than 70 %. On the basis of the physiological, biochemical and chemotaxonomic characteristics, strain G4(T) should be classified as the type strain of a novel species of the genus Agrococcus, for which the name Agrococcus carbonis sp. nov. is proposed. The type strain is G4(T) ( = MTCC 10213(T)  = DSM 22965(T)).

Pontibacter rhizosphera sp. nov., isolated from rhizosphere soil of an Indian medicinal plant Nerium indicum.

A gram-negative, motile, straight to curved rod shaped, pink pigmented bacterium was isolated from a soil sample collected from the rhizosphere of an Indian medicinal plant, Nerium indicum (Chuvanna arali) and subjected to a detailed polyphasic taxonomic study. The strain, designated as IMTB-1969(T), matched with most of the phenotypic and chemotaxonomic properties of the genus Pontibacter and represents a novel species. The major fatty acids of the strain were monounsaturated iso/anteiso branched C17 fatty acids (45.1%) and iso-C15:0 (16.5%). MK-7 was the predominant isoprenoid quinone. According to 16S rRNA gene sequence analysis, strain IMTB-1969(T) was indicated to belonged to the phylum Bacteroidetes and further phylogenetic analysis revealed that the strain IMTB-1969(T) belongs to the family Cytophagaceae and genus Pontibacter. The highest 16S rRNA gene sequence similarity was with Pontibacter korlensis CCTCC AB 206081(T) (97.2%) and lower sequence similarity was observed with other species in the genus Pontibacter (95.9-94.0%). DNA-DNA relatedness study of the strain IMTB-1969(T) confirmed that it represents a novel species. The G+C content of the genomic DNA was 52.2 (±0.5) mol%. The results of physiological and biochemical tests allowed the genotypic and phenotypic distinction of strain IMTB-1969(T) from its closest phylogenetic relatives. The strain IMTB-1969(T) should be classified as novel species of the genus Pontibacter, for which the name Pontibacter rhizosphera sp. nov. is proposed. The type strain is IMTB-1969(T) (=MTCC 10673(T) = DSM 24399(T)).

Description of a novel actinobacterium Kocuria assamensis sp. nov., isolated from a water sample collected from the river Brahmaputra, Assam, India.

A Gram-positive, pale yellow pigmented actinobacterium, strain S9-65(T) was isolated from a water sample collected from the river Brahmaputra, Assam, India and subjected to a polyphasic taxonomic study. The physiological and biochemical properties, major fatty acids (anteiso-C15:0 and anteiso-C17:0), estimated DNA G+C content (69.2 mol %) and 16S rRNA gene sequence analysis showed that strain S9-65(T) belonged to the genus Kocuria. Strain S9-65(T) exhibited highest 16S rRNA gene sequence similarity with Kocuria palustris (99.1%); however, the DNA-DNA relatedness value between strain S9-65(T) and K. palustris was 20.6%. On the basis of differential phenotypic characteristics and genotypic distinctiveness, strain S9-65(T) should be classified as representative of a novel species Kocuria, for which the name Kocuria assamensis is proposed. The type strain is S9-65(T) (=MTCC 10622(T) = DSM 23999(T)).

Description of a novel actinobacterium Microbacterium assamensis sp. nov., isolated from water sample collected from the river Brahmaputra, Assam, India.

A Gram-positive, yellow pigmented actinobacterium, strain S2-48(T) was isolated from water sample collected from the river Brahmaputra, Assam, India and subjected to a polyphasic taxonomic study. Most of the physiological and biochemical properties, major fatty acids (C(15:0) Anteiso, iso C(16:0) and C(17:0) Anteiso), estimated DNA G+C content (70.2 mol%) and 16S rRNA gene sequence analysis showed that strain S2-48(T) belonged to the genus Microbacterium. Strain S2-48(T) exhibited highest 16S rRNA gene sequence similarity with Microbacterium testaceum (97.0%); however, the DNA-DNA relatedness value between strain S2-48(T) and M. testaceum was 9.1%. On the basis of differential phenotypic characteristics and genotypic distinctiveness, strain S2-48(T) should be classified within the genus Microbacterium as a novel species, for which the name Microbacterium assamensis is proposed. The type strain is S2-48(T) (=MTCC 10486(T) = DSM 23998(T)).

Cohnella ferri sp. nov. a novel member of the genus Cohnella isolated from haematite ore.

The taxonomic position of a Gram-positive, endo-spore forming bacterium isolated from a haematite ore sample was analyzed by a polyphasic approach. The strain designated as HIO-4(T) matched most of the phenotypic and chemical characteristics of the genus Cohnella and represents a novel species. The sequence of the almost complete 16S rRNA (1489 bases) was compared with those of previously studied Cohnella type strains and confirmed that the strain belongs to the genus Cohnella. Strain HIO-4(T) differs from all other species of Cohnella by at least 3.9% at the 16S rRNA level and the moderately related species are Cohnella phaseoli (96.1%) and Cohnella yongneupensis (96.1%), respectively. Predominant polar lipids are diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE); few unknown phospholipids, mannose containing lipid, aminophospholipid and aminophosphoglycolipids. The results of physiological and biochemical tests allowed the genotypic and phenotypic distinctiveness of strain HIO-4(T) with its phylogenetic relatives and suggest that the strain HIO-4(T) should be recognized as a novel species, for which the name Cohnella ferri sp. nov. is proposed. The type strain is HIO-4(T) (=MTCC 8365(T) = JCM 16139(T)).

Production, purification and characterization of raw starch hydrolyzing thermostable acidic α-amylase from hot springs, India.

Alpha-amylase is an important hydrolytic enzyme used for various industrial processes. In the present study, Geobacillus bacterium (K1C), producing a thermostable α-amylase was isolated from Manikaran hot springs, India. We have purified and characterized the biochemical properties of α-amylase. The optimum temperature and pH for α-amylase activity was 80 °C and pH 6.0 respectively. The far-UV CD spectra of the enzyme indicated the presence of random coil conformation and showed an intermediate phase during temperature-induced unfolding. In the presence of substrate, thermostability of the α-amylase was increased as 50% initial activity was retained at 70 °C for 6 h and at 80 °C for 2 h. Moreover, the enzyme also showed remarkable pH stability as 90% of the initial activity was retained even after 48 h of incubation at pH 5.0, 6.0 and 7.0. Interestingly, amylase activity of the purified enzyme was Ca2+independent, whereas the complete inhibition of activity was observed in the presence of Cu2+, Pb2+, and Hg2+. The purified α-amylase was stable in the presence of detergents, organic solvents and Proteinase K. Furthermore, it exhibited the ability to hydrolyze raw starches (e.g. rice, wheat, corn, potato) efficiently; thus this enzyme has the potential to be used for industrial applications.

Visualizing the history of living spaces.

The technology available to building designers now makes it possible to monitor buildings on a very large scale. Video cameras and motion sensors are commonplace in practically every office space, and are slowly making their way into living spaces. The application of such technologies, in particular video cameras, while improving security, also violates privacy. On the other hand, motion sensors, while being privacy-conscious, typically do not provide enough information for a human operator to maintain the same degree of awareness about the space that can be achieved by using video cameras. We propose a novel approach in which we use a large number of simple motion sensors and a small set of video cameras to monitor a large office space. In our system we deployed 215 motion sensors and six video cameras to monitor the 3,000-square-meter office space occupied by 80 people for a period of about one year. The main problem in operating such systems is finding a way to present this highly multidimensional data, which includes both spatial and temporal components, to a human operator to allow browsing and searching recorded data in an efficient and intuitive way. In this paper we present our experiences and the solutions that we have developed in the course of our work on the system. We consider this work to be the first step in helping designers and managers of building systems gain access to information about occupants' behavior in the context of an entire building in a way that is only minimally intrusive to the occupants' privacy.