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[Substrate specificity of lipase from Pseudomonas fluorescens].

Substrate specificity of lipase isolated from the culture liquid filtrate of Pseudomonas fluorescens BKM-B-1151 was investigated with respect to vegetable oils and animal fats (olive, sunflower, cotton, mustard and soybean oils; beef and hog fats and their glycerides and fatty acid esters). The preparation showed a high specificity to the quantitative composition of the reaction mixture (substrate: enzyme ratio), chemical structure of the substrate, and the emulgator type (gelatine, gum arabic and Triton X-100). The lipase preparation hydrolyzed oils and water-insoluble fatty acid esters. The latter indicated an involvement of lipase.

Kinetics↗
PubMed1993

Substrate specificity of the heparin lyases from Flavobacterium heparinum.

A detailed knowledge about the substrate specificities of the heparin lyases is necessary when using these enzymes as tools for elucidating the sequence of heparin and heparan sulfate. The substrate specificity of heparin lyases I, II, and III have been profiled with structurally defined, heparin-derived oligosaccharides. The primary substrate specificities of heparin lyases I and III require the presence of 2-O-sulfated alpha-L-idopyranosyluronic acid and beta-D-glucopyranosyluronic acid residues, respectively, at the linkages being cleaved. Heparin lyase II demonstrates an intriguingly broad primary specificity for oligosaccharides, acting at linkages containing alpha-L-idopyranosyluronic and beta-D-glucopyranosyluronic acid as well as at linkages containing alpha-L-galactopyranosyluronic acid residues. In addition to their primary specificities, each lyase also demonstrates secondary specificities under forcing conditions. Differences in the sulfation pattern within uronic acid residues and sulfation of adjacent residues has profound impact on the ease of lyase cleavage of a glycosidic linkage. Specifically, heparin lyases I and III exhibit secondary specificity for oligosaccharides containing an unsulfated alpha-L-idopyranosyluronic acid residue. The lack of sulfation on residues adjacent to the linkage undergoing cleavage increases the action of heparin lyase III on a glycosidic linkage. In contrast, reduced sulfation on adjacent residues make glycosidic linkage resistant to heparin lyase I. The primary and secondary specificity can be rationalized on the basis of most favorable solution conformation of the uronic acid residues.

Carbohydrate Conformation↗
PubMed1980

Phospholipase A2 activity and substrate specificity of snake venom presynaptic toxins.

Beta-Neurotoxins from certain snake venoms are highly specific toxins acting at the presynaptic side of the neuromuscular junction. In this study biochemical aspects of this high specificity have been investigated. When toxins (notexin and Naja nigricollis basic phospholipase) act on a mixture of subcellular fractions obtained from brain cortex (synaptosomes, myelin, and mitochondria), the synaptosomal fraction is preferentially attacked and shows the highest release of membrane protein. As seen from isolated fractions, however, even the mitochondria are rapidly and strongly attached. Examining the phospholipase A2 activity of the toxin instead of the release of proteins reveals that synaptosomes represent the best substrate. In contrast to nonneurotoxic phospholipases A2, that from neurotoxin preferentially uses synaptosomal phosphatidylcholine as a substrate when pure phospholipids isolated from subcellular fractions are used. A relationship between the cholesterol/phospholipid ratio and the sensitivity to toxin action in the various subcellular fractions was found. These data suggest that the neurotoxic effect is mainly due to the substrate specificity of the beta-neurotoxins. It is suggested that synaptosomal phosphatidylcholine, embedded in a membrane containing a low amount of cholesterol, is a highly specific substrate for beta-neurotoxins.

Animals↗
PubMed2002

Domain closure, substrate specificity and catalysis of D-lactate dehydrogenase from Lactobacillus bulgaricus.

NAD-dependent Lactobacillus bulgaricus D-Lactate dehydrogenase (D-LDHb) catalyses the reversible conversion of pyruvate into D-lactate. Crystals of D-LDHb complexed with NADH were grown and X-ray data collected to 2.2 A. The structure of D-LDHb was solved by molecular replacement using the dimeric Lactobacillus helveticus D-LDH as a model and was refined to an R-factor of 20.7%. The two subunits of the enzyme display strong asymmetry due to different crystal environments. The opening angles of the two catalytic domains with respect to the core coenzyme binding domains differ by 16 degrees. Subunit A is in an "open" conformation typical for a dehydrogenase apo enzyme and subunit B is "closed". The NADH-binding site in subunit A is only 30% occupied, while in subunit B it is fully occupied and there is a sulphate ion in the substrate-binding pocket. A pyruvate molecule has been modelled in the active site and its orientation is in agreement with existing kinetic and structural data. On domain closure, a cluster of hydrophobic residues packs tightly around the methyl group of the modelled pyruvate molecule. At least three residues from this cluster govern the substrate specificity. Substrate binding itself contributes to the stabilisation of domain closure and activation of the enzyme. In pyruvate reduction, D-LDH can adapt another protonated residue, a lysine residue, to accomplish the role of the acid catalyst His296. Required lowering of the lysine pK(a) value is explained on the basis of the H296K mutant structure.

Binding Sites↗
PubMed1995

Modulation of the substrate specificity of the mammalian phosphatidylinositol 3-kinase by cholesterol sulfate and sulfatide.

The substrate specificity of the purified, mammalian phosphatidylinositol 3-kinase is subject to modulation by detergents, which are able to switch substrate specificity in vitro in favor of PtdInsP2. This effect of the detergents is due to an activation of the phosphatidylinositol biphosphate 3-kinase activity, while the phosphatidylinositol 3-kinase activity is inhibited. The selective inhibition of the phosphatidylinositol 3-kinase activity (p110 alpha/p85 alpha) is shown here also to be observed by employing cholesterol sulfate or sulfatide at low micromolar concentrations, whereas cholesterol and androsterone sulfate fail to inhibit. These naturally occurring sulfated lipids have at these concentrations no effect on the phosphatidylinositol bisphosphate 3-kinase activity but inhibit the manganese-dependent intrinsic protein kinase activity, thus switching substrate specificity toward the more highly phosphorylated inositol lipids. Cholesterol sulfate and sulfatide inhibit the free catalytic subunit p110 alpha but fail to inhibit the homologous phosphatidylinositol 3-kinase from Saccharomyces cerevisiae (Vps34p), suggesting that these sulfated lipids act specifically on the mammalian phosphatidylinositol 3-kinase. Consistent with this specificity, the regulatory subunit (p85), which is not conserved in the yeast enzyme, is found to play an important role for the affinity of these inhibitors. The implications for the phosphatidylinositol 3-kinase activity in vivo are discussed.

Adenosine Triphosphate↗
PubMed2005

Identification of conserved prolyl residue important for transport activity and the substrate specificity range of yeast plasma membrane Na+/H+ antiporters.

Yeast plasma membrane Na+/H+ antiporters are divided according to their substrate specificity in two distinct subfamilies. To identify amino acid residues responsible for substrate specificity determination (recognition of K+), the Zygosaccharomyces rouxii Sod2-22 antiporter (non-transporting K+) was mutagenized and a collection of ZrSod2-22 mutants that improved the KCl tolerance of a salt-sensitive Saccharomyces cerevisiae strain was isolated. Several independent ZrSod2-22 mutated alleles contained the replacement of a highly conserved proline 145 with a residue containing a hydroxyl group (Ser, Thr). Site-directed mutagenesis of Pro145 proved that an amino acid with a hydroxyl group at this position is enough to enable ZrSod2-22p to transport K+. Simultaneously, the P145(S/T) mutation decreased the antiporter transport activity for both Na+ and Li+. Replacement of Pro145 with glycine resulted in a ZrSod2-22p with extremely low activity only for Na+, and the exchange of a charged residue (Asp, Lys) for Pro145 completely stopped the activity. Mutagenesis of the corresponding proline in the S. cerevisiae Nha1 antiporter (Pro146) confirmed that this proline of the fifth transmembrane domain is a critical residue for antiporter function. This is the first evidence that a non-polar amino acid residue is important for the substrate specificity and activity of yeast Nha antiporters.

Alleles↗
PubMed2005

Towards a better understanding of the substrate specificity of the UDP-N-acetylglucosamine C4 epimerase WbpP.

WbpP is the only genuine UDP-GlcNAc (UDP-N-acetylglucosamine) C4 epimerase for which both biochemical and structural data are available. This represents a golden opportunity to elucidate the molecular basis for its specificity for N-acetylated substrates. Based on the comparison of the substrate binding site of WbpP with that of other C4 epimerases that convert preferentially non-acetylated substrates, or that are able to convert both acetylated and non-acetylated substrates equally well, specific residues of WbpP were mutated, and the substrate specificity of the mutants was determined by direct biochemical assays and kinetic analyses. Most of the mutations tested were anticipated to trigger a significant switch in substrate specificity, mostly towards a preference for non-acetylated substrates. However, only one of the mutations (A209H) had the expected effect, and most others resulted in enhanced specificity of WbpP for N-acetylated substrates (Q201E, G102K, Q201E/G102K, A209N and S143A). One mutation (S144K) totally abolished enzyme activity. These data indicate that, although all residues targeted in the present study turned out to be important for catalysis, determinants of substrate specificity are not confined to the substrate-binding pocket and that longer range interactions are essential in allowing proper positioning of various ligands in the binding pocket. Hence prediction or engineering of substrate specificity solely based on sequence analysis, or even on modelling of the binding pocket, might lead to incorrect functional assignments.

Acetylation↗
PubMed1994

Identification of substrate specificity determinants for the cell cycle-regulated NIMA protein kinase.

NIMA is a cell cycle-regulated protein kinase required for the G2/M transition in the filamentous fungus Aspergillus nidulans. Previous biochemical characterization of the recombinant enzyme indicated that NIMA is a protein serine/threonine specific kinase with beta-casein being the best substrate from the many proteins and peptides tested (Lu, K.P., Osmani, S.A., and Means, A.R. (1993) J. Biol. Chem. 268, 8769-8776). However, substrate specificity or physiologically relevant substrates for NIMA remained unknown. In search for a peptide substrate for this enzyme, we screened an assembled library of synthetic peptides that each contained a phosphorylation site for a known protein kinase and found an excellent peptide substrate for NIMA, phospholemman 42-72 (PLM(42-72)). NIMA kinase phosphorylated PLM(42-72) uniquely and stoichiometrically on Ser63 with a Vmax of 1.4 mumol/min/mg and apparent Km of 20.0 microM. These kinetic constants were about 10-fold higher and 3-fold lower than those for beta-casein, respectively. A detailed analysis of substrate specificity determinants using synthetic peptide analogs of PLM(42-72) indicated that Phe-Arg-Xaa-Ser/Thr represents the optimal primary sequence for NIMA kinase phosphorylation. Replacement of the Arg at P-2 with Ala resulted in a 6-fold increase in Km and 2-fold decrease in Vmax, while substitution of the Phe at P-3 with Ala abolished NIMA phosphorylation. These results reveal the unique nature of substrate recognition by the NIMA kinase and should prove valuable in the search for biologically relevant NIMA substrates.

Amino Acid Sequence↗
PubMed1987

Substrate specificity of the carbon monoxide-dependent cytochrome P-450 kinetics.

The substrate-dependent kinetics of the carbon monoxide-inhibited cytochrome P-450 activity and its light reversibility is reinvestigated in microsomal preparations. In order to find out whether the substrate specificity is mediated by an isoenzyme-specific binding of carbon monoxide with different dissociation constants an experimental design has been chosen where it could be established that essentially the same isoenzyme component was involved in two different monooxygenase reactions, i.e., the O-dealkylation of 7-ethoxycoumarin and the 7-hydroxylation of coumarin. The dissociation constant kD(CO) of the ferrous cytochrome P-450 carbon monoxide complex is 6-fold higher in the presence of 7-ethoxycoumarin than in the presence of coumarin. But the light-induced relative changes of the Warburg partition coefficient for the 7-ethoxycoumarin deethylation and for coumarin 7-hydroxylation do not differ remarkably from each other. These relative changes are shown to represent the ratio of the photoinduced rate constant to the spontaneous rate constant of the dissociation for the ferrous cytochrome P-450 carbon monoxide complex. The differences in the dissociation constants are assigned to substrate specific effects on the carbon monoxide binding, indicating a substrate-specific change of the free binding enthalpy for carbon monoxide.

Animals↗
PubMed1999

Substrate specificity of the periplasmic dipeptide-binding protein from Escherichia coli: experimental basis for the design of peptide prodrugs.

Pure dipeptide-binding protein (DppA) from Escherichia coli was studied in a filter binding assay to determine its binding specificity. A substrate:DppA stoichiometry of 1:1 was found with both [14C]AlaAla and Ala[14C]Phe. Surprisingly, substrate binding did not vary over the pH range pH 3-9.5. Different dipeptides yielded liganded protein with various pI values, implying that DppA can undergo subtly different conformational changes to accommodate different substrates. Using [125I]Tyr-peptides as substrates in competition assays, the relative binding affinities for a range of dipeptides were found to parallel their overall transport rates into E. coli through the dipeptide permease (Dpp), showing that DppA alone controls the specificity of Dpp. With a series of substituted glycyl peptides, binding affinity was progressively enhanced by alkylation (with methyl to butyl) of the N-terminal alpha-amino group. Thus, results from this approach provide an essential experimental basis, which complements the information from the crystal structure of DppA, for the design of peptidomimetic antibacterials targeted for transport through Dpp.

Amino Acids, Branched-Chain↗
PubMed2002

The SPOT technique as a tool for studying protein tyrosine phosphatase substrate specificities.

The activity of protein tyrosine phosphatases (PTPs) is restricted by their substrate specificities. The analysis of PTP specificity was greatly helped by the discovery that "substrate-trapping" PTP mutants, such as PTP-1B D181A, stably and specifically bind their substrates. We have set up a PTP substrate specificity assay based on the SPOT technique, which involves the microsynthesis of (phospho)peptides on membranes. To validate this approach, substrate trapping PTP-1B was tested on its cognate ligand, the autophosphorylated insulin receptor (IR). On SPOT membranes, IR peptides with phosphotyrosine 1163 were efficiently bound by PTP1B D181A, and dephosphorylated by PTP-1B. Phosphotyrosine 1163 was preferred over the neighboring 1158 and 1162 phosphotyrosines. PTP-1B also recognized IR-like motifs in Trk autophosphorylation domains, and STAT 5 phosphopeptides. Using a gridded 20-by-20 SPOT library, we show that peptides with the YZM motif (Z: phosphotyrosine) are the strongest ligands for PTP-1B D181A, but not the optimal substrates for dephosphorylation by wild-type PTP1B. In addition we show that PTP-1B and PTP-beta dephosphorylation efficiency is strongly modulated by the introduction of phospho-serine or phospho-threonine in their cognate phospho-tyrosine substrates. Altogether our data illustrate that the SPOT technique is a highly efficient tool for the study of PTP substrate specificity.

Amino Acid Motifs↗
PubMed1997

Substrate specificity of human O6-methylguanine-DNA methyltransferase for O6-benzylguanine derivatives in oligodeoxynucleotides.

To investigate the substrate specificity of human O6-methylguanine-DNA methyltransferase (MGMT) for O6-benzylguanine (6BG) derivatives incorporated in oligodeoxynucleotides, we prepared 25-mer lengths of sequences containing various 6BG derivatives and their related compounds and then measured the ability of these derivatives to inactivate MGMT in vitro. Oligodeoxynucleotides containing a 6BG, O6-(2-fluorobenzyl)guanine (2F-6BG), O6-(3-fluorobenzyl)guanine (3F-6BG), O6-(4-fluorobenzyl)guanine (4F-6BG), O6-benzylhypoxanthine (6BH), or O6-methylguanine (6MG) were all good substrates for MGMT, and no obvious differences were observed among them. Oligodeoxynucleotides containing N2-isobutyrylated 6BG and 6MG showed only a slightly reduced capacity for inactivating MGMT compared to N2-nonmodified forms of these derivatives. No obvious differences were observed in the corresponding double-stranded and single-stranded oligodeoxynucleotides. MGMT substrate specificity for the 6BG derivatives in the oligodeoxynucleotide was found to be quite different from that seen in our previous study [Mineura, K., et al. (1994) Int. J. Cancer 58, 706-712; (1995) Int. J. Cancer 63, 148-151. Kohda, K., et al. (1995) Biol. Pharm. Bull. 18, 424-430] and others [Moschel, R. C., et al. (1992) J. Med. Chem. 35, 4486-4491. Chae, M.Y., et al. (1994) J. Med. Chem. 37, 342-347] using the corresponding free bases. In brief, (i) 6BG, 3F-6BG, and 4F-6BG greatly inhibited human MGMT, whereas 2F-6BG, 6BH, and 6MG displayed much weaker activity; (ii) any modifications at the 2-amino group of the 6BG resulted in severe reductions in the ability to inactivate MGMT. These results obtained by the experiments using oligodeoxynucleotides and free bases suggest that human MGMT has low substrate specificity for 6BGs in oligodeoxynucleotides. Conformational changes in human MGMT which favor binding to oligodeoxynucleotides containing 6BG derivatives and the subsequent transfer of their benzyl groups may account for the difference in substrate specificity between the incorporated 6BG derivatives and their free base form.

Antineoplastic Agents↗
PubMed2005

Substrate specificity and novel selective inhibitors of TNF-alpha converting enzyme (TACE) from two-dimensional substrate mapping.

We report a systematic analysis of the P1' and P2' substrate specificity of TNF-alpha converting enzyme (TACE) using a peptide library and a novel analytical method, and we use the substrate specificity information to design novel reverse hydroxamate inhibitors. Initial truncation studies, using the amino acid sequence around the cleavage site in precursor-TNF-alpha, showed that good turnover was obtained with the peptide DNP-LAQAVRSS-NH2. Based on this result, 1000 different peptide substrates of the form Biotin-LAQA-P1'-P2'-SSK(DNP)-NH2 were prepared, with 50 different natural and unnatural amino acids at P1' in combination with 20 different amino acids at P2'. The peptides were pooled, treated with purified microsomal TACE, and the reaction mixtures were passed over a streptavidin affinity column to remove unreacted substrate and the N-terminal biotinylated product. C-terminal cleavage products not binding to streptavidin were subjected to liquid chromatography/mass spectrometry analysis where individual products were identified and semiquantitated. 25 of the substrates were resynthesized as discrete peptides and assayed with recombinant TACE. The experiments show that recombinant TACE prefers lipophilic amino acids at the P1' position, such as phenylglycine, homophenylalanine, leucine and valine. At the P2' position, TACE can accommodate basic amino acids, such as arginine and lysine, as well as certain non-basic amino acids such as citrulline, methionine sulfoxide and threonine. These substrate preferences were used in the design of novel reverse hydroxamate TACE inhibitors with phenethyl and 5-methyl-thiophene-methyl side-chains at P1', and threonine and nitro-arginine at P2'.

ADAM Proteins↗
PubMed1983

Human platelet phenolsulphotransferase M and P: substrate specificities and correlation with in vivo sulphoconjugation of paracetamol and salicylamide.

Human platelet phenolsulphotransferase exists in two functional forms. M and P. In this study the substrate specificity of the two forms has been further delineated by correlating activities in different individuals with various substrates. m-Tyramine, noradrenaline, adrenaline, 5-hydroxytryptamine, p-hydroxyamphetamine, isoprenaline, salbutamol and l-naphthol were all specific substrates for the M form of the enzyme. Paracetamol, a mixed substrate, was predominantly metabolized by the M form. Salicylamide at 5 microM was a substrate for the P form but became and M substrate at higher concentration. Phenol itself, a specific substrate for phenolsulphotransferase P at 10 microM, also became an M substrate at 1 mM concentration. These substrate specificities were confirmed with the selective inhibitor, dichloronitrophenol. In this study, we measured phenolsulphotransferase activity in platelets from 13 individuals selected on the basis of their wide variation in ability to sulphoconjugate paracetamol and salicylamide in vivo. There was no significant relationship between the in vivo pattern with either drug and the activity of platelet phenolsulphotransferase assayed with paracetamol or salicylamide respectively.

Acetaminophen↗
PubMed1981

Substrate specificity of CTP-synthetase from E. coli.

The substrate specificity of CTP-synthetase from E. coli was investigated by means of UTP analogs. This study revealed that the three main structural elements of the UTP molecule were important for the substrate specificity of the enzyme. CTP-synthetase seems to possess an absolute requirement for the beta-D-ribose 5-triphosphate part in UTP. Substitutents in 5-position of UTP, exceeding the size of a tritium atom abolish substrate function.

Carbon-Nitrogen Ligases↗
PubMed1985

Substrate specificity of beta-collagenase from Clostridium histolyticum.

The substrate specificity of beta-collagenase from Clostridium histolyticum has been investigated by measuring the rate of hydrolysis of more than 50 tri-, tetra-, penta-, and hexapeptides covering the P3 to P3' subsites of the substrate. The choice of peptides was patterned after sequences found in the alpha 1 and alpha 2 chains of type I collagen. Each peptide contained either a 2-furanacryloyl (FA) or cinnamoyl (CN) group in subsite P2 or the 4-nitrophenylalanine (Nph) residue in subsite P1. Hydrolysis of the P1-P1' bond produces an absorbance change in these chromophoric peptides that has been used to quantitate the rates of their hydrolysis under first order conditions ([S] much less than KM) from kcat/KM values have been obtained. The identity of the amino acids in all six subsites (P3-P3') markedly influences the hydrolysis rates. In general, the best substrates have Gly in subsites P3 and P1', Pro or Ala in subsite P2', and Hyp, Arg, or Ala in subsite P3'. This corresponds well with the frequency of occurrence of these residues in the Gly-X-Y triplets of collagen. In contrast, the most rapidly hydrolyzed substrates do not have residues from collagen-like sequences in subsites P2 and P1. For example, CN-Nph-Gly-Pro-Ala is the best known substrate for beta-collagenase with a kcat/KM value of 4.4 X 10(7) M-1 min-1, in spite of the fact that there is neither Pro nor Ala in P2 or Hyp nor Ala in P1. These results indicate that the previously established rules for the substrate specificity of the enzyme require modification.

Amino Acid Sequence↗
PubMed2004

Mutation of active site residues in the chitin-binding domain ChBDChiA1 from chitinase A1 of Bacillus circulans alters substrate specificity: use of a green fluorescent protein binding assay.

A fluorescent binding assay was developed to investigate the effects of mutagenesis on the binding affinity and substrate specificity of the chitin-binding domain of chitinase A1 from Bacillus circulans WL-12. The chitin-binding domain was genetically fused to the N-terminus of a green fluorescent protein, and the polyhistidine-tagged hybrid protein was expressed in Escherichia coli. Residues likely to be involved in the binding site were mutated and their contributions to binding and substrate specificity were evaluated by affinity electrophoresis and depletion assays. The experimental binding isotherms were analyzed by non-linear regression using a modified Langmuir equation. Non-conservative substitution of tryptophan residue (W687) nearly abolished chitin-binding affinity and dramatically lowered chitosan binding while retaining the original level of curdlan binding. Double mutation E668K/P689A had altered specificity for several substrates and also impaired chitin binding significantly. Other substitutions in the binding site altered substrate specificity but had little effect on overall affinity for chitin. Interestingly, mutation T682A led to a higher specificity towards chitinous substrates than the wildtype. Furthermore, the ChBD-GFP hybrid protein was tested for use in diagnostic staining of cell walls of fungi and yeast and for the detection of fungal infections in tissue samples.

Amino Acid Sequence↗
PubMed1993

Dipeptidyl peptidase IV (DPP-IV) from pig kidney cleaves analogs of bovine growth hormone-releasing factor (bGRF) modified at position 2 with Ser, Thr or Val. Extended DPP-IV substrate specificity?

The literature reported DPP-IV substrate specificity includes oligopeptides with a penultimate Pro, Hyp or Ala residue. Bovine GRF is a substrate for DPP-IV and is rapidly degraded by the enzyme via removal of its N-terminal Tyr-Ala. Incubation of selected GRF analogs from the [X2,Ala15,Leu27]bGRF(1-29)NH2 series with a porcine-kidney-derived DPP-IV in PBS (pH 7.4) resulted in cleavage at the X2-Asp3 bond. The extent of enzymatic hydrolysis varied with X2 as reflected in the following relative cleavage rates: Ala2 (100%), Ser2 (4%), Thr2 (2.5%), Val2 (0.53%), Ile2 (0%). These cleavages were sequestered when similar experiments were performed in the presence of the DPP-IV-specific inhibitor N-epsilon-(p-NO2-benzyloxycarbonyl)-Lys-Pro-OH. A side reaction, buffer-induced deamidation of Asn8, contributed less than 5% of the total substrate degradation. Although our finding qualitatively extends the DPP-IV substrate specificity to also include N-terminal X-Ser, X-Thr and X-Val sequences, quantitatively, relatively fast cleavages of the GRFs with Ala2 make the latter preferred substrates for DPP-IV. The data presented here indicates that the observed GRF(3-29) fragment formation upon incubation of Ser2- and Thr2-substituted bGRF analogs in bovine plasma could have been DPP-IV-related.

Animals↗