Thermostable Peptide Research Articles

Design of a Novel Peptide with Esterolytic Activity toward PET by Mimicking the Catalytic Motif of Serine Hydrolases.

Serine hydrolases have become increasingly important for recycling PET plastics. However, their properties are inherently constrained by their 3D structure, which in turn limits the conditions for their application. Considering peptides as catalysts for industrial depolymerization processes can help us to escape some of these limitations. In this article, a 25 amino acid thermostable peptide, HSH-25, was designed to depolymerize PET. The peptide incorporates a His-Ser-His motif, inspired by the catalytic triad found in the serine hydrolase family, into a β-hairpin fold. Stability of the fold was investigated by molecular dynamics simulations. Esterolytic activity of the peptide toward model substrates was detected within a pH range from pH 7 to pH 9.5. Degradation of polymeric PET substrates was confirmed by atomic force microscopy imaging on spin-coated PET thin films.

Isolation and Characterization of Bacteriocin Producing Levilactobacillus brevis Strain ABRIINW-K from Buffalo Dung

Bacteriocins are proteins secreted by many species of bacteria to inhibit other bacteria, thus eliminating competitors to gain resources. Bacteria from the Lactobacillus group are known for their applications as probiotics and food preservatives. They have earned a reputation for producing substances that inhibit the growth of other microorganisms, which include organic acids, diacetyl, and bacteriocins. Produced by the ribosomes, bacteriocins are cationic proteins that inhibit other bacteria coexisting within a shared ecological habitat. Due to their potential uses in a variety of applications large-scale production of Bacteriocins would be necessary. The study aimed to identify and characterize Lactobacillus bacteria that produce potent bacteriocins and to analyze the antimicrobial activity and stability of the isolated bacteriocin under various physical and biochemical conditions. A total of 50 samples including buffalo dung, cheese, and rhizospheric region of plants were screened to isolate 8 Lactobacillus Li-1, Li-2, Li-3, Li-4, Li-5, Li-6, Li-7, and Li-8, confirmed by gram staining and other biochemical tests. The cell free supernatant from the Li-3 strain showed higher inhibition of Escherichia coli and Staphylococcus aureus, as compared to the other isolated strains. Li-3 strain was further identified as Levilactobacillus brevis strain ABRIINW-K by 16S rRNA gene sequencing. The bacteriocin isolated from this strain is a thermostable peptide (~6kDa), which is characteristic of class II bacteriocins, with potent antibacterial activity against Lactobacillus rhamnosus, Escherichia coli, and Salmonella enterica.

Characterization of inhibitory supernatants produced by bacteria isolated from goat milk

Several compounds can be produced by lactic acid bacteria (LAB) isolated from milk such as bacteriocins, small cationic and thermostable peptides, that possess antimicrobial activity. The technological application of these antimicrobial compounds became important for the industry mainly because of chemical preservatives. This study aimed to isolate LAB with anti-Listeria monocytogenes activity from raw goat milk and to characterize their inhibitory cell-free supernatants (CFS). It was performed Isolation of LAB from raw goat milk, inhibitory activity of the CFS, characterization of CFS inhibitory activity under different conditions, mode of action of the active CFS, adsorption of the anti-Listeria substances present in the CFS of LS2 to L. monocytogenes, molecular identification of LAB isolates with anti-Listeria activity. Three isolated strains produced CFS with active inhibitory substances of proteinaceous nature, identified by 16S rDNA as Weissella cibaria (LS1) and Lactococcus lactis (LS2 and LS3). The CFS of LS2 showed the highest anti-Listeria activity (1600 arbitrary units/mL) after 2 h at pH 6.0 and was bacteriostatic and active at low pH at temperatures of 4 °C to 80 °C. The results suggest that the LS2 strain is a biopreservative culture with potential for applications to control L. monocytogenes in foods.

Purification and Identification of Novel Antioxidant Peptides Isolated from Geoffroea decorticans Seeds with Anticoagulant Activity.

Geoffroea decorticans is a xerophilous deciduous tree present in most arid forests of southern South America, which is commonly used in traditional medicine. The seeds of this tree have been previously investigated for their singular chemical composition, but their protein content has been poorly investigated. Herein, we report the isolation, purification, and characterization of a set of thermostable peptides derived from Geoffroea decorticans seeds (GdAPs) with strong antioxidant and anticoagulant activities. The most potent antioxidant peptides showed a half maximal inhibitory concentration (IC50) of 35.5 ± 0.3 µg/mL determined by 1,1-diphenyl-2-picrylhydrazyl (DPPH). They also caused a dose-dependent prolongation of the aPTT clotting time with an IC50 value of ~82 µg/mL. Interestingly, MALDI-TOF/MS analysis showed the presence of three major peptides with low molecular weights of 2257.199 Da, 2717.165 Da, and 5422.002 Da. The derived amino-acid sequence of GdAPs revealed their unique structural features, exhibiting homology with various proteins present in the genome of Arachis hypogaea. All in all, our data suggest a direct applicability of GdAPs for pharmaceutical purposes.

BING, a novel antimicrobial peptide isolated from Japanese medaka plasma, targets bacterial envelope stress response by suppressing cpxR expression

Antimicrobial peptides (AMPs) have emerged as a promising alternative to small molecule antibiotics. Although AMPs have previously been isolated in many organisms, efforts on the systematic identification of AMPs in fish have been lagging. Here, we collected peptides from the plasma of medaka (Oryzias latipes) fish. By using mass spectrometry, 6399 unique sequences were identified from the isolated peptides, among which 430 peptides were bioinformatically predicted to be potential AMPs. One of them, a thermostable 13-residue peptide named BING, shows a broad-spectrum toxicity against pathogenic bacteria including drug-resistant strains, at concentrations that presented relatively low toxicity to mammalian cell lines and medaka. Proteomic analysis indicated that BING treatment induced a deregulation of periplasmic peptidyl-prolyl isomerases in gram-negative bacteria. We observed that BING reduced the RNA level of cpxR, an upstream regulator of envelope stress responses. cpxR is known to play a crucial role in the development of antimicrobial resistance, including the regulation of genes involved in drug efflux. BING downregulated the expression of efflux pump components mexB, mexY and oprM in P. aeruginosa and significantly synergised the toxicity of antibiotics towards these bacteria. In addition, exposure to sublethal doses of BING delayed the development of antibiotic resistance. To our knowledge, BING is the first AMP shown to suppress cpxR expression in Gram-negative bacteria. This discovery highlights the cpxR pathway as a potential antimicrobial target.

Antimicrobial activity of bacteriocin‐producing Carnobacterium spp. isolated from healthy Patagonian trout and their potential for use in aquaculture

In this study, bacteria from the intestines of Patagonian trout displaying fish pathogen antimicrobial activity were isolated and characterized. Isolates T4, T15 and M5 showed extracellular antimicrobial activity against the fish pathogen Carnobacterium maltaromaticum CECT 4020, by the agar well diffusion assay. They were identified as Gram-positive catalase-negative bacilli belonging to the genus Carnobacterium (based on 99.93% identity with the 16S rRNA gene sequence of C. maltaromaticum DSM 20342T). After 48 hr of incubation in MRS broth, the highest antimicrobial activity titre (327,680 AU/ml) was recorded for T4, followed by T15 and M5. The antimicrobial agents produced by these isolates were thermostable peptides. LC-MS/MS peptide analysis indicated that T4 and T5 bacteriocins belonged to the class IIa. The identified peptides for T4 bacteriocin included almost the entire divercin V41 sequence, which was previously believed to be exclusively produced by Carnobacterium divergens. In addition, no negative fish health effects were observed after administrating T4 to zebrafish. Among the isolates from healthy Patagonian trout, due to its early bacteriocin production, its greater inhibition titre and its innocuous feature, Carnobacterium sp. T4 emerged as the most promising isolate for in vivo challenges targeting its further aquaculture application.

Rapid synthesis and decoration of reduced graphene oxide with gold nanoparticles by thermostable peptides for memory device and photothermal applications

This article presents novel, rapid, and environmentally benign synthesis method for one-step reduction and decoration of graphene oxide with gold nanoparticles (NAuNPs) by using thermostable antimicrobial nisin peptides to form a gold-nanoparticles-reduced graphene oxide (NAu-rGO) nanocomposite. The formed composite material was characterized by UV/Vis spectroscopy, X-ray diffraction, Raman spectroscopy, X-ray photoelectron spectroscopy, field emission scanning electron microscopy, and high-resolution transmission electron microscopy (HR-TEM). HR-TEM analysis revealed the formation of spherical AuNPs of 5–30 nm in size on reduced graphene oxide (rGO) nanosheets. A non-volatile-memory device was prepared based on a solution-processed ZnO thin-film transistor fabricated by inserting the NAu-rGO nanocomposite in the gate dielectric stack as a charge trapping medium. The transfer characteristic of the ZnO thin-film transistor memory device showed large clockwise hysteresis behaviour because of charge carrier trapping in the NAu-rGO nanocomposite. Under positive and negative bias conditions, clear positive and negative threshold voltage shifts occurred, which were attributed to charge carrier trapping and de-trapping in the ZnO/NAu-rGO/SiO2 structure. Also, the photothermal effect of the NAu-rGO nanocomposites on MCF7 breast cancer cells caused inhibition of ~80% cells after irradiation with infrared light (0.5 W cm−2) for 5 min.

Protease inhibitor in scorpion (Mesobuthus eupeus) venom prolongs the biological activities of the crude venom

It is hypothesized that protease inhibitors play an essential role in survival of venomous animals through protecting peptide/protein toxins from degradation by proteases in their prey or predators. However, the biological function of protease inhibitors in scorpion venoms remains unknown. In the present study, a trypsin inhibitor was purified and characterized from the venom of scorpion Mesobuthus eupeus, which enhanced the biological activities of crude venom components in mice when injected in combination with crude venom. This protease inhibitor, named MeKTT-1, belonged to Kunitz-type toxins subfamily. Native MeKTT-1 selectively inhibited trypsin with a Kivalue of 130 nmol·L−1. Furthermore, MeKTT-1 was shown to be a thermo-stable peptide. In animal behavioral tests, MeKTT-1 prolonged the pain behavior induced by scorpion crude venom, suggesting that protease inhibitors in scorpion venom inhibited proteases and protect the functionally important peptide/protein toxins from degradation, consequently keeping them active longer. In conclusion, this was the first experimental evidence about the natural existence of serine protease inhibitor in the venom of scorpion Mesobuthus eupeus, which preserved the activity of venom components, suggests that scorpions may use protease inhibitors for survival.

Highly fluorescent peptide nanoribbon impregnated with Sn-porphyrin as a potent DNA sensor

Highly fluorescent and thermo-stable peptide nanoribbons (PNRs) were fabricated by solvothermal self-assembly of a single peptide (D,D-diphenyl alanine peptides) with Sn-porphyrin (trans-dihydroxo[5,10,15,20-tetrakis(p-tolyl)porphyrinato] Sn(IV) (SnTTP(OH)2)). The structural characterization of the as-prepared nanoribbons was performed by transmitting electron microscopy (TEM), scanning electron microscopy (SEM) and atomic force microscopy (AFM), FT-IR and Raman spectroscopy, indicating that the lipophilic Sn-porphyrins are impregnated into the porous surface formed in the process of nanoribbon formation through intermolecular hydrogen bonding of the peptide main chains. Consequently the Sn-porphyrin-impregnated peptide nanoribbons (Sn-porphyrin-PNRs) exhibited typical UV-visible absorption spectrum of the monomer porphyrin with a red shifted Q-band, and their fluorescence quantum yield was observed to be enhanced compared to that of free Sn-porphyrin. Interestingly the fluorescence intensity and lifetimes of Sn-porphyrin-PNRs were selectively affected upon interaction with nucleotide base sequences of DNA while those of free Sn-porphyrins were not affected by binding with any of the DNA studied, indicating that DNA-induced changes in the fluorescence properties of Sn-porphyrin-PNRs are due to interaction between DNA and the PNR scaffold. These results imply that Sn-porphyrin-PNR will be useful as a potent fluorescent protein analogue and as a biocompatible DNA sensor.

Identification, Purification and Characterization of Laterosporulin, a Novel Bacteriocin Produced by Brevibacillus sp. Strain GI-9

BackgroundBacteriocins are antimicrobial peptides that are produced by bacteria as a defense mechanism in complex environments. Identification and characterization of novel bacteriocins in novel strains of bacteria is one of the important fields in bacteriology.Methodology/FindingsThe strain GI-9 was identified as Brevibacillus sp. by 16 S rRNA gene sequence analysis. The bacteriocin produced by strain GI-9, namely, laterosporulin was purified from supernatant of the culture grown under optimal conditions using hydrophobic interaction chromatography and reverse-phase HPLC. The bacteriocin was active against a wide range of Gram-positive and Gram-negative bacteria. MALDI-TOF experiments determined the precise molecular mass of the peptide to be of 5.6 kDa and N-terminal sequencing of the thermo-stable peptide revealed low similarity with existing antimicrobial peptides. The putative open reading frame (ORF) encoding laterosporulin and its surrounding genomic region was fished out from the draft genome sequence of GI-9. Sequence analysis of the putative bacteriocin gene did not show significant similarity to any reported bacteriocin producing genes in database.ConclusionsWe have identified a bacteriocin producing strain GI-9, belonging to the genus Brevibacillus sp. Biochemical and genomic characterization of laterosporulin suggests it as a novel bacteriocin with broad spectrum antibacterial activity.

Mechanism and Kinetics of Peptide Partitioning into Membranes from All-Atom Simulations of Thermostable Peptides

Partitioning properties of transmembrane (TM) polypeptide segments directly determine membrane protein folding, stability, and function, and their understanding is vital for rational design of membrane active peptides. However, direct determination of water-to-bilayer transfer of TM peptides has proved difficult. Experimentally, sufficiently hydrophobic peptides tend to aggregate, while atomistic computer simulations at physiological temperatures cannot yet reach the long time scales required to capture partitioning. Elevating temperatures to accelerate the dynamics has been avoided, as this was thought to lead to rapid denaturing. However, we show here that model TM peptides (WALP) are exceptionally thermostable. Circular dichroism experiments reveal that the peptides remain inserted into the lipid bilayer and are fully helical, even at 90 degrees C. At these temperatures, sampling is approximately 50-500 times faster, sufficient to directly simulate spontaneous partitioning at atomic resolution. A folded insertion pathway is observed, consistent with three-stage partitioning theory. Elevated temperature simulation ensembles further allow the direct calculation of the insertion kinetics, which is found to be first-order for all systems. Insertion barriers are DeltaH(in)(double dagger) = 15 kcal/mol for a general hydrophobic peptide and approximately 23 kcal/mol for the tryptophan-flanked WALP peptides. The corresponding insertion times at room temperature range from 8.5 mus to 163 ms. High-temperature simulations of experimentally validated thermostable systems suggest a new avenue for systematic exploration of peptide partitioning properties.

Proteomic characterization of the thermostable toxins from Naja naja venom

Naja naja snake venom presents abundant thermostable peptides. Many of them possess useful pharmacological activity that may be employed for drug development. For the proteomic characterization of such toxins, in the present study, Naja naja venom solution was heated up to 100°C for 10, 30, 60, 120, 180 and 300 minutes and protein fractions of non-heated and heated venom were analyzed by two-dimensional nano-liquid chromatography coupled online with tandem mass spectrometry. After heating for 300 minutes, a total of 32 peptides were still detected in the supernatant. The identified peptides belong to the following groups: cardiotoxins, neurotoxins and cytotoxins. It was found that thermostable peptides are able to preserve their analgesic activity after a long heating time in formalin test. Mice injected with 15 μg/g of 60-minute heated venom or with 25 μg/g of 300-minute heated venom revealed even a better analgesic activity than those treated with lidocaine.

Differential elicitation of an aspartic protease inhibitor: Regulation of endogenous protease and initial events in germination in seeds of Vigna radiata

Plant aspartic proteases are of recent origin with their physiological significance in crucial processes emerging. Reports on the significance of aspartic protease inhibitors and their endogenous proteases in seeds of plants are scanty. This paper reports the purification of an aspartic protease inhibitor from the seeds of Vigna radiata, its control of the endogenous aspartic protease and their subsequent role in the early germination events. The role of the aspartic protease inhibitor and the enzyme in initial stages of germination of V. radiata has been tracked by differential timed expression and germination assays. The expression pattern revealed maximum expression of the inhibitor in the dormant seeds while the enzyme was predominant in the germinating seeds. Their expression patterns and interactions indicate their significance in initiation of germination. The expression of other classes of proteases was monitored during germination and a model predicting the events occurring during proteolysis of the storage protein in germination is hypothesized. The inhibitor was a linear, hydrophobic, pH stable and thermostable peptide with molecular weight of 1660 Da. The purified inhibitor showed a p I of 4.36 with the sequence as AEIYN KDGNK LDLYG. The inhibitor was found to be stable in a broad range of pH from 2 to 10 with an optimum of 3.0. The half-life of VrAPI at 100 °C was 30 min whereas the maximum activity was observed at 37 °C. The initial kinetic analysis of the inhibitor against the endogenous protease showed an IC 50 value of 11 nM while the value of the inhibition rate constant K i was 34 × 10 −9 M.

Molecular cloning, structural analysis and modelling of the AcAFP antifungal peptide from Aspergillus clavatus

An abundantly secreted thermostable peptide (designed AcAFP) with a molecular mass of 5777 Da was isolated and purified in a previous work from a local strain of A. clavatus (VR1). Based on the N-terminal amino acid (aa) sequence of the AcAFP peptide, an oligonucleotide probe was derived and allowed the amplification of the encoding cDNA by RT-PCR. This cDNA fragment encodes a pre-pro-protein of 94 aa which appears to be processed to a mature product of 51 aa cys-rich protein. The deduced aa sequence of the pre-pro-sequence reveals high similarity with ascomycetes antifungal peptide. Comparison of the nucleotide sequence of the genomic fragment and the cDNA clone revealed the presence of an open reading frame of 282 bp interrupted by two small introns of 89 and 56 bp with conserved splice site. The three-dimensional (3D) structure modeling of AcAFP exhibits a compact structure consisting of five anti-parallel β barrel stabilized by four internal disulfide bridges. The folding pattern revealed also a cationic site and spatially adjacent hydrophobic stretch. The antifungal mechanism was investigated by transmission and confocal microscopy. AcAFP cause cell wall altering in a dose-dependent manner against the phytopathogenic fungus Fusarium oxysporum.

Cloning and Expression of Antibacterial Goat Lactoferricin from Escherichia coli AD494(DE3)pLysS Expression System

Goat lactoferricin (GLfcin), an antibacterial peptide, is released from the N terminus of goat lactoferrin by pepsin digestion. Two GLfcin-related cDNAs, GLfcin L and GLfcin S, encoding Ala20-Ser60 and Ser36-Ser60 of goat lactoferrin, respectively, were cloned into the pET-23a(+) expression vector upstream from (His)6-Tag gene and transformed into Escherichia coli AD494(DE3)pLysS expression host. After being induced by isopropyl-β-d -thiogalactopyranoside (IPTG), two (His)6-Tag fused recombinant lactoferricins, GLfcin L-His·Tag and GLfcin S-His·Tag, were expressed in soluble form within the E. coli cytoplasm. The GLfcin L-His·Tag and GLfcin S-His·Tag were purified using HisTrap affinity chromatography. According to an antibacterial activity assay using the agar diffusion method, GLfcin L-His·Tag had antibacterial activity against E. coli BCRC 11549, Staphylococcus aureus BCRC 25923, and Propionibacterium acnes BCRC 10723, while GLfcin S-His·Tag was able to inhibit the growth of E. coli BCRC 11549 and P. acnes BCRC 10723. These two recombinant lactoferricins behaved as thermostable peptides, which could retain their activity for up to 30 min of exposure at 100°C.

Protein kinase inhibitor peptide (PKI): A family of endogenous neuropeptides that modulate neuronal cAMP-dependent protein kinase function

Signal transduction cascades involving cAMP-dependent protein kinase are highly conserved among a wide variety of organisms. Given the universal nature of this enzyme it is not surprising that cAMP-dependent protein kinase plays a critical role in numerous cellular processes. This is particularly evident in the nervous system where cAMP-dependent protein kinase is involved in neurotransmitter release, gene transcription, and synaptic plasticity. Protein kinase inhibitor peptide (PKI) is an endogenous thermostable peptide that modulates cAMP-dependent protein kinase function. PKI contains two distinct functional domains within its amino acid sequence that allow it to: (1) potently and specifically inhibit the activity of the free catalytic subunit of cAMP-dependent protein kinase and (2) export the free catalytic subunit of cAMP-dependent protein kinase from the nucleus. Three distinct PKI isoforms (PKIalpha, PKIbeta, PKIgamma) have been identified and each isoform is expressed in the brain. PKI modulates neuronal synaptic activity, while PKI also is involved in morphogenesis and symmetrical left-right axis formation. In addition, PKI also plays a role in regulating gene expression induced by cAMP-dependent protein kinase. Future studies should identify novel physiological functions for endogenous PKI both in the nervous system and throughout the body. Most interesting will be the determination whether functional differences exist between individual PKI isoforms which is an intriguing possibility since these isoforms exhibit: (1) cell-type specific tissue expression patterns, (2) different potencies for the inhibition of cAMP-dependent protein kinase activity, and (3) expression patterns that are hormonally, developmentally and cell-cycle regulated. Finally, synthetic peptide analogs of endogenous PKI will continue to be invaluable tools that are used to elucidate the role of cAMP-dependent protein kinase in a variety of cellular processes throughout the nervous system and the rest of the body.

Purification and primary structure of low molecular mass peptides from scorpion ( Buthus sindicus) venom

The primary structures of four low molecular mass peptides (Bs 6, 8, 10 and 14) from scorpion Buthus sindicus were elucidated via combination of Edman degradation and matrix-assisted laser desorption ionization mass spectrometry. Bs 8 and 14 are cysteine-rich, thermostable peptides composed of 35–36 residues with molecular weights of 3.7 and 3.4 kDa, respectively. These peptides show close sequence homologies (55–78%) with other scorpion chlorotoxin-like short-chain neurotoxins (SCNs) containing four intramolecular disulfide bridges. Despite the sequence variation between these two peptides (37% heterogeneity) their general structural organization is very similar as shown by their clearly related circular dichroism spectra. Furthermore, Bs6 is a minor component, composed of 38 residues (4.1 kDa) containing six half-cystine residues and having close sequence identities (40–80%) with charybdotoxin-like SCNs containing three disulfide bridges. The non-cysteinic, bacic and thermolabile Bs10 is composed of 34 amino acid residues (3.7 kDa), and belongs to a new class of peptides, with no sequence resemblance to any other so far reported sequence isolated from scorpions. Surprisingly, Bs10 shows some limited sequence analogy with oocyte zinc finger proteins. Results of these studies are discussed with respect to their structural similarities within the scorpion LCNs, SCNs and other biologically active peptides.

A new peptide (1150Da) selectively activates the calcium-calmodulin sensitive isoform of cyclic nucleotide phosphodiesterase from human myometrium

The cyclic nucleotide phosphodiesterase enzymatic system is examined in extracts of human myometrium and four individual phosphodiesterase isoforms have been isolated and characterized. A new thermostable peptide, recently purified in rat and calf myometrium, is able to stimulate up to 55-fold, the calcium-calmodulin dependent phosphodiesterase isoform. Activation of cAMP hydrolysis is by far the most marked with a 55-fold maximal stimulation at a concentration of 0.1μM peptide and a IC50 value estimated at 30nM. For cGMP hydrolysis, the maximal effect (×25) obtained at 40nM peptide is lesser and the IC50 value is in the 10nM range. Furthermore, we verified that classical calmodulin antagonists such as calmidazolium or trifluoroperazine did not change stimulation of the calcium-calmodulin phosphodiesterase by the peptide, indicating that the myometrial peptide is different from calmodulin. To our knowledge, this is the first evidence for such a strong and selective stimulation of one isoform of the phosphodiesterase enzymatic system by a natural peptide.

An express immunological method for detection of human seminal plasma

Monoclonal antibodies (Mabs) against human seminal plasma (HSP) were produced and during screening procedures dissociation constants of the antigen/antibody complexes were determined. Mab 1E5 was selected for further studies because of its high reactivity in an enzyme-linked immunoassay (ELISA) and high affinity for its corresponding antigen. The specificity of Mab 1E5 was checked in absorption ELISA with human organ extracts and some biological secretions. It was established that the 1E5-corresponding epitope was a thermostable peptide moiety which could be detected in HSP, only. This monoclonal antibody was used for the development of an express method for detection of human semen. The assay was applied for screening of 57 cases of suspected rape. A complete correlation was found between the results obtained by the proposed test and by routine microscopic methods. The newly designed immunoassay is reliable, it is easily performed and it is less time-consuming.

Characterization of a thermostable LH-immunoreactive material from bovine pituitary gland

1. 1. A thermostable peptide, immunologically similar to the β-subunit of ovine lutropin, was isolated from a heated extract of bovine pituitary glands. 2. 2. The relative molecular mass of the peptide as well as its amino acid composition differed significantly from the lutropin subunits. 3. 3. Antibodies were raised to a preparation of this peptide.