Wort Protein Research Articles

Malt quality profile of barley predicted by near‐infrared spectroscopy using partial least squares, Bayesian regression, and artificial neural network models

AbstractDue to the significant cost and time involved in identifying barley lines with superior malting quality, the malting industry is searching for accurate and rapid methods to expedite the selection of superior barley lines that meet breeder's goals. The aim of this study is to compare partial least squares regression (PLSR) with advanced statistical models (Bayesian and machine learning) and reliably assess their performance in predicting malt quality traits from near infra‐red (NIR) spectral data using barley grains. Using spectral data as predictors and the malt quality traits as references, PLSR outperformed Bayesian and PCA‐ANN models for diastatic power (DP), alpha amylase (AA), malt extract (ME), wort protein (WP), soluble to total protein (S/T) ratio, and free amino nitrogen (FAN). WP had the best prediction performance for all models, with the best‐performing model, PLSR, having (RPD) values of 0.55 (1.5). The influential wavelength regions identified based on the variable importance in projection (VIP) scores and coefficient estimates for PLSR and Bayesian models, respectively, were comparatively similar for all malt quality traits. Based on these findings, PLSR analysis and wavelength selection techniques would enhance the future design and optimization of NIR prediction models in malt quality improvement programs.

Addition of Hop (Humulus Lupulus L.) Bitter Acids Yields Modification of Malt Protein Aggregate Profiles during Wort Boiling.

Hop bitter acids are used in the brewing industry to give beer bitterness. However, much of this bitterness is lost during processing, specifically during the wort boiling step. One of the major causes might be the interaction with protein-protein complexes. Therefore, the aim of this study was to clarify the role of hop bitter acids in protein aggregate formation using a proteomic approach. The effect of hop addition on protein composition was analyzed by liquid chromatography-mass spectrometry/MS (LC-MS/MS), and further analyses were performed to characterize the wort before and after boiling. Addition of hop bitter acids yielded a change in wort protein profiles, and hop bitter acids were found to bind primarily to less abundant proteins which are not related to beer quality traits, such as foam or haze. Wort protein aggregate profiles were revealed, and findings from this study suggested the precipitation of particular proteins in the aggregates during boiling when hops were added.

Experimental and numerical investigation of beer foam

The frothy foam on top of beer is produced by bubbles of gas, predominantly carbon dioxide, rising to the surface. The chemical components that produce the head are wort protein, yeast, and hop residue. This involves a large number of chemical components and numerous physical interactions. The beer foam and, especially, its stability is an essential quality characteristic of a beer. A consumer defines a beer's head by its stability, quantity, lacing (glass adhesion or cling), whiteness, creaminess, and strength. To achieve an accurate prediction of beer foam formation and collapse is challenging because complex numerical models are required to account for these nonlinear beer foam effects. To analyze a new design of a beer bottom-to-top pouring system for the startup company Einstein 1, we first set up the experimental tests of this pouring system. Afterward, we performed the associated repeatability studies to achieve stable working conditions. To study beer foam formation and its collapse, we employed a multiphase Reynolds-averaged Navier–Stokes solver that considered two inter-penetrating continua, which allowed accounting for multi-component phases and mass and heat transfer between these phases. We numerically and experimentally investigated beer foam patterns, beer heights, beer/foam ratio, foam height, foam stability, and foam volume fractions. We performed grid sensitivity studies and validated the numerical solver by comparing results against model test measurements. The results indicated that that the higher the temperature of the beer and the higher the tap pressure is, the greater the foam development and the associated foam height are but not necessarily the foam stability.

Modification mechanism of protein in rice adjuncts upon extrusion and its effects on nitrogen conversion during mashing

The traditional production of wort with adjunct-introduced was achieved by double mashing procedure, which hindered the utilization of proteins in adjunct and led to a deficiency of nitrogen in wort. In this study, the modification mechanism of the extrusion pretreatment on the structure characterization of rice flour protein was investigated. The decoction mashing procedure was performed to enhance the nitrogen conversion of the extruded rice adjunct. Decreased solubility along with disrupted secondary and tertiary structures of rice protein were observed after extrusion. As a result, the total nitrogen, free amino nitrogen, and free amino acids content of wort with extruded rice adjunct-introduced were improved by 23.28 %, 34.67 %, and 7.33 %, respectively, which could be verified by the electrophoretic patterns of the wort protein. The application of extrusion as a pretreatment of adjuncts can promote the protein availability of adjuncts in the decoction mashing stage.

Interaction of proteins and amino acids with iso-\u03b1-acids during wort preparation in the brewhouse

This paper investigates the binding behavior of iso-α-acids from hops on free wort amino acids and proteins concerning the wort production process in breweries. The studies were carried out with different amino acids, bovine serum albumin and wort. To identify the nature of reaction between iso-α-acids and these substances, analyses of free amino nitrogen, HPLC and isothermal titration calorimetry were performed. According to the results, the iso-α-acids do not form covalent bonds with free amino acids of wort. However, iso-α-acids, especially isohumulone and isoadhumulone, form ionic bonds with wort proteins. A distinction must be made between proteins that are present in the hot trub, and those that are still dissolved in the hot wort. Proteins that are already coagulated and precipitated no longer react with iso-α-acids. Future experiments will investigate whether the established ionic bonds between iso-α-acids and proteins from the wort preparation process are maintained during fermentation until the finished beer or beer foam. If this is the case, which is induced by the experiments, there is a measurable loss of iso-α-acids in the hot wort, but at the same time, a gain for the later beer foam retention, as the iso-α-acids will stabilize it.

On the Use of Bovine Serum Albumin for the Spectrophotometric Determination of Protein in Unhopped Wort

Malt soluble protein (SP) can be determined by different analytical procedures such as Kjeldahl, combustion, or spectrophotometry. From a practical standpoint, the spectrophotometric method is the most preferred. This method requires a linear calibration with different malts in a wide SP range. For this calibration however, protein in wort is determined by Kjeldahl as the reference method. Considering that some soluble protein standards could be used as an alternative for the Kjeldahl analysis in the calibration step, the objective of this research was to evaluate the use of bovine serum albumin (BSA) as a protein standard in the determination of SP. Results showed that the BSA standard curve overestimates protein in wort and, therefore, SP values. However, estimated vs. actual wort protein values were highly associated (R2=0.9984) and the correspondent regression lines had similar slopes. Taking this into account, the BSA standard curve was adjusted with wort protein values observed in international malt check samples (SP range= 4.3–6.4% of malt, dry basis). The resultant equation from the BSA adjusted curve [wort protein, %wt/vol= (1.4007* A215-225) − 0.0071] improved the estimation of protein in wort by minimizing differences in magnitude.

Vesicular structures formed from barley wort proteins and iso-humulone

In beer, the main amphiphilic components are protein and iso-humulone. Two major populations of protein are identified as lipid transfer protein (LTP) (9.7 kDa) and protein Z (43 kDa). In this paper, protein and iso-humulone are extracted from barley malt and hop, respectively, based on the brewing process. Mixtures of protein and iso-humulone are mixed at different concentrations and centrifuged. Supernatants are analyzed by asymmetric flow field-flow fractionation (AF4) coupled to UV with multi-angle light scattering (MALS) detection as well as by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE). The presence of aggregates and their structures are investigated by light microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The results show that protein and iso-humulone can form aggregates from molecular level by AF4-UV-MALS and SDS-PAGE. The interaction is shown as solution depletion which is analyzed by AF4-UV-MALS. With 300 mg/L iso-humulone and 3 g/L protein in bulk, the decrease of protein peak levels off. The peak of protein Z is preferentially decreased as an effect of iso-humulone being present, suggesting that interaction between these populations is favored. The iso-humulone/protein aggregates consist of both undefined irregular aggregates as well as spherical aggregates. The spherical aggregates are observed in light microscopy, SEM and TEM. From SEM, it is clear that there are two types of spherical aggregates: rough and smooth. With TEM it can be observed that the smooth aggregates consist of a thin layer of the aggregated proteins and iso-humulone enclosing a liquid domain. This structure can best be described as an iso-humulone/protein vesicle. The rough vesicles are formed by further precipitation at the surface of the smooth vesicles.

Phenotypic and allelic variation for wort protein Z in Australian and Canadian barleys

Protein Z is a major component in beer foam. Two-dimensional electrophoresis was used to analyze wort proteins of two Australian (Buloke and Commander) and two Canadian (CDC Meredith and Bentley) varieties. The Canadian barley contained more abundant proteins from MW 40–45 kDa (pI 5 to 7). These proteins were identified as either protein Z4 or protein Z7 using liquid chromatography–mass spectrometry. Full-length gene of protein Z4 and Z7 were sequenced from Canadian and Australian barleys. Sequence differences were identified in the coding region and upstream regions of the two genes, resulting in protein sequence and expression variations. Molecular markers were designed according to the indels in the upstream regions of protein Z4 and Z7 genes. These markers were highly correlated to wort protein Z content in Canadian and Australian varieties. The Canadian barleys contained ‘high level’ genotypes for protein Z4 and Z7 while most Australian barleys had ‘low level’ genotypes for protein Z4, Z7 or both. The markers identified in this study provide a valuable tool for the selection of protein Z alleles in marker-assisted breeding. Total protein Z content was assessed using different steeping conditions, and increasing air-rest time increased protein Z content in 15 varieties.

Interfacial properties and interaction between beer wort protein fractions and iso-humulone

It is well known that both protein and iso-humulone, originating from barley malt and hops respectively, play important roles in beer foam stability. In this study, we hypothesize that the interaction of iso-humulone and different beer proteins will give rise to adsorbed layers with different characteristics at hydrophobic surfaces. The properties of these adsorbed layers are likely to play a crucial role for beer foam formation and stability.Beer protein and iso-humulone were extracted based on the brewing process from barley malt and hop respectively. The proteins were fractionated into two fractions (lipid transfer protein (LTP) and a mixed fraction of LTP and protein Z) and their interfacial properties as well as interaction with iso-humulone in solution and at hydrophobic surfaces were investigated by surface rheology and ellipsometry. The results show that the composition and properties of the adsorbed layers depend on how the layer is formed and that iso-humulone plays an important role for the characteristics of the adsorbed protein layers. The interaction between iso-humulone and protein Z is found to have pronounced effects of yielding a coherent and elastic adsorbed layer and, thus, for beer foam stability.

Evolution of a quality testing program for improving malting barley in Canada

Edney, M. J., MacLeod, A. L. and LaBerge, D. E. 2014. Evolution of a quality testing program for improving malting barley in Canada. Can. J. Plant Sci. 94: 535–544. The quality of Canadian malting barley has continually improved since malt barley was exported in the late 1800s. Improvements were linked to a dynamic evaluation system that evolved with a better understanding of malting biochemistry and as suitable methods were developed. Methods became more accurate and more specific in their ability to define quality. They progressed from sensory evaluation, to surmising malt quality from barley protein levels, to the first micro-maltings followed by automated laboratory-scale maltings. Malt quality analysis started simply with malt extract and diastatic power followed by wort protein. As the necessity for cell wall breakdown became better understood, analyses like wort viscosity, fine/coarse grind extract differences and wort β-glucan were adopted. A continuum of cultivars were released in Canada, based on this evaluation system, starting with the six-rowed releases OAC 21, then Montcalm and Bonanza, followed by the two-rowed releases Betzes, Klages, Harrington and AC Metcalfe. Release of future cultivars will depend on an evolving evaluation system that could include; barley homogeneity, specific starch-degrading enzymes, individual amino acids and specific traits such as low lipoxygenase and low phytic acid barley. The result will be development and release of cultivars with better defined quality that can fill specialized niches in the malting and brewing industries of the future.

Quantitative trait loci of barley malting quality trait components in the Stellar/01Ab8219 mapping population

Malting barley is of high economic and scientific importance. Determining barley grains that are suitable for malting involves measuring malting quality, which is an expensive and complex process. In order to decrease the cost of phenotyping and accelerate the process of developing superior malting barley cultivars, markers for marker-assisted breeding are needed. In this study, we identified quantitative trait loci (QTLs) for malting traits in a Stellar/01Ab8219 F6:8 recombinant inbred line population grown at Aberdeen and Tetonia, Idaho, USA in 2009 and 2010. We identified QTLs associated with malt extract (ME), wort protein, soluble/total protein (S/T), diastatic power (DP), alpha-amylase, beta-glucan (BG) and free amino nitrogen (FAN) at a logarithm of odds score ≥2.5 using a high-density genetic map produced by merging Diversity Arrays Technology markers with the current single nucleotide polymorphism map. Novel QTLs were identified for DP and FAN on chromosome 5H, S/T on 6H, and BG and ME on 7H. Dissection of the genetic regions associated with malting traits suggests the involvement of multiple molecular pathways. The resulting molecular markers may prove useful for barley improvement.

Антиоксиданты из растительного сырья в технологии стабилизации пива

Through every stage of the brewing process, oxygen badly affects the taste and stability of the final beer. at the stage of boiling wort with hop oxygen performs two different functions. It is positive for isomerization process of α-acids, and it is negative for degradation processes in isoα-acids. as iso-α-acids are destructed analytical value of wort and beer bitterness reduces, and its nature deteriorates. Normal alcoholic fermentation happens in the absence of oxygen, and this is the cornerstone of the technological scheme of accelerated production of beer. Every researcher recognized the negative role that oxygen plays in maturation. oxidation can be stopped or slowed down by using antioxidants which either react with generated peroxide free radicals or destroy peroxides without free radicals being generated. To prevent oxidative degradation of iso-α-acids, accumulation dynamics has been studied with regard to trends of accumulation of bitter substances in the hopped wort depending on the time when antioxidants from the bark of oak and peppermint leaves are added into boiling wort and hop mixture. Studies have shown that it is desirable to add antioxidants from the bark of oak and peppermint leaves in 50 minutes since after wort with hop is getting boiled, in order to prevent oxidative destruction of iso-α-acids. It is found that antioxidant phenolic compounds from oak bark and leaves of peppermint most effectively contribute to the coagulation of wort proteins.

The impact of different ale brewer’s yeast strains on the proteome of immature beer

BackgroundIt is well known that brewer’s yeast affects the taste and aroma of beer. However, the influence of brewer’s yeast on the protein composition of beer is currently unknown. In this study, changes of the proteome of immature beer, i.e. beer that has not been matured after fermentation, by ale brewer’s yeast strains with different abilities to degrade fermentable sugars were investigated.ResultsBeers were fermented from standard hopped wort (13° Plato) using two ale brewer’s yeast (Saccharomyces cerevisiae) strains with different attenuation degrees. Both immature beers had the same alcohol and protein concentrations. Immature beer and unfermented wort proteins were analysed by 2-DE and compared in order to determine protein changes arising from fermentation. Distinct protein spots in the beer and wort proteomes were identified using Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and MS/MS and revealed common beer proteins, such as lipid transfer proteins (LTP1 and LTP2), protein Z and amylase-protease inhibitors. During fermentation, two protein spots, corresponding to LTP2, disappeared, while three protein spots were exclusively found in beer. These three proteins, all derived from yeast, were identified as cell wall associated proteins, that is Exg1 (an exo-β-1,3-glucanase), Bgl2 (an endo-β-1,2-glucanase), and Uth1 (a cell wall biogenesis protein).ConclusionYeast strain dependent changes in the immature beer proteome were identified, i.e. Bgl2 was present in beer brewed with KVL011, while lacking in WLP001 beer.

Proteome analysis of the wort boiling process

A comprehensive proteome map was constructed of brewers wort. The map consisted of protein identification on two-dimensional gel electrophoresis (2DE) images and identified 63 out of 202 protein spots, which were categorized into 20 protein species. To analyze the modification of protein Z during wort boiling, protein Z spots on 2DE gel of the sweet wort, the boiled wort and the trub were analyzed by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI TOF-MS) and then the spectra were compared. The analysis identified several specific signals detected only in the trub, suggesting that specific modification occurred in the precipitated protein Z during wort boiling. The protein Z spot on the 2DE gel of the precipitate was further analyzed by liquid chromatography mass spectrometry/mass spectrometry. The analysis identified low molecular weight fragment (1.3kDa) derived from wound induced protein (barwin) in the protein Z spot of the trub. These results suggested that protein Z was precipitated by binding with comparatively small size specific fragment(s) derived from sweet wort protein, i.e., barwin during wort boiling. Our results and understandings have application for quality assurance and control in commercial brewing practice, and development of DNA markers for malting barley breeding.

Comparison of beer quality attributes between beers brewed with 100% barley malt and 100% barley raw material

Brewing with 100% barley using the Ondea® Pro exogenous brewing enzyme product was compared to brewing with 100% barley. The use of barley, rather than malt, in the brewing process and the consequences for selected beer quality attributes (foam formation, colloidal stability and filterability, sensory differences, protein content and composition) was considered. The quality attributes of barley, malt, kettle-full-wort, cold wort, unfiltered beer and filtered beer were assessed. A particular focus was given to monitoring changes in the barley protein composition during the brewing process and how the exogenous OndeaPro® enzymes influenced wort protein composition. All analyses were based on standard brewing methods described in ASBC, EBC or MEBAK. To monitor the protein changes two-dimensional polyacrylamide gel electrophoresis was used. It was shown that by brewing beer with 100% barley and an appropriate addition of exogenous Ondea® Pro enzymes it was possible to efficiently brew beer of a satisfactory quality. The production of beers brewed with 100% barley resulted in good process efficiency (lautering and filtration) and to a final product whose sensory quality was described as light, with little body and mouthfeel, very good foam stability and similar organoleptic qualities compared to conventional malt beer. In spite of the sensory evaluation differences could still be seen in protein content and composition.

INVESTIGATION OF HORDEINS DURING BREWING AND THEIR INFLUENCE ON BEER HAZE BY PROTEOME ANALYSIS

ABSTRACT Colloidal storage haze is one of the principal indicators of beer quality. In this study, the changes of malt hordeins during the brewing process and the roles of malt hordeins in the formation of beer haze were investigated using two-dimensional electrophoresis (2-DE). It was found that most of malt hordeins disappeared during the brewing process and only the few faint hordein spots (about eight spots) were detected in beer haze. Moreover, these spots were found in wort protein and beer haze protein. The complex 2-DE patterns of malt hordein, wort and beer haze displayed a relationship with some chemical modifications during the brewing. These spots were analyzed and identified as B and γ3 hordein and storage protein (Hordeum vulgare) by mass spectrometry. Our results suggested that hordein might be minor haze-active proteins in beer but was a critical factor in the formation of beer haze. PRACTICAL APPLICATION The formation of haze is a serious quality problem that limits the storage life of beer products, an issue that has become increasingly important to brewers. It is conventionally accepted that proline-rich hordeins play a major role in the formation of haze. In this study, it was found that most of the hordeins disappeared and that small amounts of B hordein and γ3 hordein from the malt in beer haze were detected by proteomics analysis. Moreover, this study demonstrated that hordeins in beer were not a key contributor to beer haze formation. To our knowledge, to date, this result in the existing literature of similar studies has not been reported. This study laid the theoretical foundation for further investigations to determine the mechanism of the formation of beer haze. An improved understanding of the impact of brewing on hordeins could potentially provide further scope for optimizing beer quality.

Tracking the Progress of Congress Mashing with Osmolyte Concentration and Malt Extract Value in North American Barley Cultivars and Relationships between Wort Osmolyte Concentration, Malt Extract Value, and ASBC Measures of Malt Quality

This study was conducted to test three hypotheses: 1) barley malt osmolyte concentration (OC) would increase more rapidly during mashing than malt extract (ME); 2) OC would better discriminate between differing barley malts during mashing than ME; and 3) OC after mashing would better correlate with ASBC measures of malt quality than ME. Malts of six two-row and six six-row barley cultivars were mashed in a micromasher, and aliquots were removed for wort OC and ME measurements at six time points during a 115-min mashing regime. For all cultivars combined, over the first 55 min of mashing wort OC values increased 120% compared with 91% for ME. At the end of the mashing regime (115 min) wort OC values further increased to 132%, whereas wort ME values remained approximately the same (91–92%). This indicates that the mass of material in wort remained approximately the same after 55 min but was further degraded to lower molecular weight compounds, increasing wort molarity and, thus, OC but not ME. Steptoe, a feed barley, was separated to a much greater degree from elite malting cultivars by wort OC than by ME values throughout the mashing regime. Except for very early in mashing (5 min), there was better separation of all cultivars by OC than by ME values throughout mashing, as determined with least significant difference analysis, indicating that OC may be better than ME for judging differences in cultivar malt quality. Linear correlations of time-course mash OC versus ME values were excellent for all cultivars combined (r = 0.977, P < 0.0001) and for two- and six-row cultivars separately. Correlations of the 115-min mashing OC and ME values with ASBC malt quality measurements were considerably better for OC than for ME (e.g., wort protein [r = 0.819, P = 0.001 for OC; r = 0.407, P = 0.189 for ME]; diastatic power [r = 0.765, P = 0.004 for OC; r = 0.287, P = 0.365 for ME]; α-amylase [r = 0.737, P = 0.006 for OC; r = 0.584, P = 0.046 for ME]; and β-glucan [r = −0.0.754, P = 0.005 for OC; r = −0.0.595, P = 0.041 for ME]). This study supports all three hypotheses.

Crystal structure of Hyp-1, a St. John’s wort protein implicated in the biosynthesis of hypericin

Hypericin, a red-colored naphtodianthrone, is a natural product synthesized in the medicinal plant Hypericum perforatum, widely known as St. John's wort. Hypericin has been attracting a growing attention of the pharmaceutical industry because of its potential application in various therapies, including the treatment of depression. In vivo, hypericin is synthesized by dimerization of emodin in a complicated multistep reaction that is reportedly catalyzed by a small (17.8kDa) protein, Hyp-1. Based on relatively low sequence similarity ( approximately 50%), Hyp-1 has been tentatively classified as a plant PR-10 (pathogenesis-related class 10) protein. Members of the PR-10 family are ubiquitous plant proteins associated with stress control and tissue differentiation but with no clearly understood molecular mechanism. They have, however, a well-defined folding canon, consisting of an extended antiparallel beta-sheet wrapped around a C-terminal alpha-helix, enclosing in the protein interior a huge cavity, in which various hydrophobic ligands can be bound. Apart from Hyp-1, only two other PR-10 members have been found to possess enzymatic activity (S-norcoclaurine synthase and TcmN aromatase/cyclase). In this paper, we report a high-resolution crystal structure of Hyp-1, confirming that it indeed has a PR-10 fold. The protein binds multiple polyethylene glycol molecules, some of which occupy the hydrophobic cavity. The crystallographic model illustrates a high degree of conformational adaptability of both interacting partners for efficient binding. We have been unable, however, to dimerize emodin to hypericin using Hyp-1 as biocatalyst. This puzzling result does not have a clear explanation at this time.

Wort Boiling by Batch Rectification--Possibilities for Reducing Required Evaporation

Today, energy costs are very high and are predicted to rise further. Because approx. 2,260 kJ is required to evaporate 1 L of wort (or its water content), it is advantageous to reduce total evaporation during wort boiling. The total evaporation required to achieve the target concentration in cast-out wort is a property of the solution and is determined by the vapor-liquid equilibrium of unwanted flavors in wort and starting concentration in the kettle-up wort. Thus, total evaporation cannot be reduced by existing wort-boiling systems, although this is often asserted. It was theorized that the only way to possibly reduce the required overall evaporation is by fractional distillation or rectification. To test this theory, a new wort-boiling system was constructed using the basics of evaporation and rectification. In this new rectification wort-boiling system, a side stream of wort is constantly drawn from the wort kettle and fed into a rectification column connected to the kettle. Thus, the evaporating vapor and recirculating wort are in direct contact, which concentrates unwanted flavors in the vapor. Because of this, the discharged steam has a concentration of unwanted flavors that is much higher than that produced by normal boiling systems. Test trials were performed with the new wort-boiling system, and its evaporation efficiency was determined. Results of the measurements showed that an equal reduction of unwanted flavors was achieved with approx. 50% less overall evaporation than that required by a normal wort-boiling system. The evaporation efficiency, thus, was doubled. Using this new system, it is possible to reduce the required overall evap�oration during batch processing of wort boiling by as much as 50%. If the boiling time is also reduced, the resulting wort has lower thermal stress and a desirable wort protein profile. The required rectification column can be added to nearly any existing wort-boiling system.

St. John's Wort protein, p27 SJ, regulates the MCP-1 promoter

St. John's Wort is commonly known for its antiviral, antidepressant, and cytotoxic properties, but traditionally St. John's Wort has also been used to treat inflammation. In this study, we sought to characterize the mechanisms used by St. John's Wort to treat inflammation by examining the effect of the recently isolated protein from St. John's Wort, p27 SJ on the expression of MCP-1. By employing an adenovirus expression vector, we demonstrate that a low concentration of p27 SJ upregulates the MCP-1 promoter through the transcription factor C/EBPβ. In addition, we found that C/EBPβ-homologous protein (CHOP) or siRNA-C/EBPβ significantly reduced the ability of p27 SJ to activate MCP-1 gene expression. Results from protein-protein interaction studies illustrate the existence of a physical interaction between p27 SJ and C/EBPβ in microglial cells. The use of chromatin immunoprecipitation assay (ChIP) led to the identification of a new cis-element that is responsive to C/EBPβ within the MCP-1 promoter. Association of C/EBPβ with MCP-1 DNA was not affected by the presence of p27 SJ. The biological activity of MCP-1 produced by cultures of adenovirus-p27 SJ transduced cells was increased relative to controls as measured by the transmigration of human Jurkat cells. Thus, we conclude that at high concentration, p27 SJ is a potential agent that may be developed as a modulator of MCP-1 leading to the inhibition of the cytokine-mediated inflammatory responses.