Lenka Zakova

Lenka Zakova
The Czech Academy of Sciences | AVCR · Institute of Organic Chemistry and Biochemistry

PhD

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72
Publications
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Introduction
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Publications

Publications (72)
Article
Preptin is a 34-amino-acid-long peptide derived from the E-domain of a precursor of insulin-like growth factor 2 (pro-IGF2) with bone-anabolic and insulin secretion amplifying properties. Here, we describe the synthesis, structures, and biological activities of six shortened analogues of human preptin. Eight- and nine-amino-acid-long peptide amides...
Article
We adapted a radioligand receptor binding assay for measuring insulin levels in unknown samples. The assay enables rapid and accurate determination of insulin concentrations in experimental samples, such as from insulin-secreting cells. The principle of the method is based on the binding competition of insulin in a measured sample with a radiolabel...
Poster
Full-text available
Design of insulin receptor B isoform selective analogue.
Article
Full-text available
The members of the insulin superfamily are well conserved across the evolution tree. We recently showed that four viruses in the Iridoviridae family possess genes that share high similarity with human insulin and IGF-1. By chemically synthesizing single chain (sc, IGF-1 like) forms of these viral insulin/IGF-1 like peptides (VILPs), we previously s...
Article
OBJECTIVE: Members of the insulin/insulin-like growth factor (IGF) superfamily are well conserved across the evolutionary tree. We recently showed that four viruses in the Iridoviridae family possess genes that encode proteins highly homologous to human insulin/IGF-1. Using chemically synthesized single-chain (sc), i.e., IGF-1-like, forms of the vi...
Article
Full-text available
Objective: Members of the insulin/insulin-like growth factor (IGF) superfamily are well conserved across the evolutionary tree. We recently showed that four viruses in the Iridoviridae family possess genes that encode proteins highly homologous to human insulin/IGF-1. Using chemically synthesized single-chain (sc), i.e., IGF-1-like, forms of the v...
Article
Full-text available
Insulin is produced and stored inside the pancreatic β-cell secretory granules, where it is assumed to form Zn ²⁺ -stabilized oligomers. However, the actual storage forms of this hormone and the impact of zinc ions on insulin production in vivo are not known. Our initial X-ray fluorescence experiment on granules from native Langerhans islets and in...
Article
Full-text available
Preparation of both 125 I-labeled insulin and insulin-like growth factor 1 (IGF-1) was critical because it enabled a detailed characterization of binding properties of these important hormones towards their cognate transmembrane receptors. Binding modes of hundreds of hormone derivatives were analyzed using competition radioligand binding assays. T...
Article
Full-text available
Insulin-like growth factors 2 and 1 (IGF2 and IGF1) and insulin are closely related hormones that are responsible for the regulation of metabolic homeostasis, development and growth of the organism. Physiological functions of insulin and IGF1 are relatively well-studied, but information about the role of IGF2 in the body is still sparse. Recent dis...
Preprint
Full-text available
Members of the insulin/IGF superfamily are well conserved across the evolutionary tree. We recently showed that four viruses in the Iridoviridae family possess genes that encode proteins highly homologous to human insulin/IGF-1. Using chemically synthesized single chain (sc), i.e. IGF-1-like, forms of the viral insulin/IGF-1 like peptides (VILPs),...
Article
Full-text available
Cone snails have evolved a variety of insulin-like molecules that may help with the development of better treatments for diabetes.
Article
A new method, pressure assisted partial filling affinity capillary electrophoresis, has been developed to study noncovalent molecular interactions of the hexamer of human insulin (HI) with biologically relevant ligands, basic phenolic neurotransmitters serotonin and dopamine, basic amino acid arginine, and very weakly acidic phenol, in alkaline aqu...
Article
Full-text available
Insulin and insulin-like growth factor 1 (IGF-1) are closely related hormones involved in the regulation of metabolism and growth. They elicit their functions through activation of tyrosine kinase-type receptors: insulin receptors (IR-A and IR-B) and IGF-1 receptor (IGF-1R). Despite similarity in primary and three-dimensional structures, insulin an...
Article
Full-text available
Insulin-like growth factors 1 and 2 (IGF-1 and -2) are protein hormones involved in normal growth and development, but also in lifespan regulation and cancer. They exert their functions mainly through the IGF-1R receptor, or by binding to the isoform A of insulin receptor (IR-A). The development of IGF-1 and IGF-2 antagonists is of great clinical i...
Article
Full-text available
Non‐natural compounds mimicking the actions of proteins and large peptides can find a plethora of applications in modulating protein‐protein interactions. In this study, we tested the biological properties of three new tripodal and trifunctional scaffolds designed for the solid‐phase synthesis of three different peptides on the same scaffold. Using...
Article
Full-text available
Human insulin-like growth factor 1 (IGF-1) is a 70-amino acid protein hormone, with key impact on growth, development and lifespan. The physiological and clinical importance of IGF-1 prompted challenging chemical and biological trials towards the development of its analogs as molecular tools for the IGF-1 receptor (IGF1-R) studies, and as new thera...
Article
Full-text available
A significant drawback of the exogenous administration of insulin to diabetics is the non-physiological profile of insulin action resulting in the insufficient suppression of hepatic glucose production, which is the main contributing factor to diabetic hyperglycemia under fasting conditions and the basis of the challenge to restore a more physiolog...
Article
Full-text available
Human insulin is a pivotal protein hormone controlling metabolism, growth and ageing, and whose malfunctioning underlies diabetes, some cancers and neuro-degeneration. Despite its central position in human physiology, the in vivo oligomeric state and conformation of insulin in its storage granules in the pancreas are not known. In contrast, many in...
Article
We designed a combinatorial library of tri-functional scaffold-derived compounds, which were derivatized with 30 different in-house-made azides. The compounds were proposed to mimic insulin receptor (IR)-binding epitopes in the insulin molecule and bind to and activate this receptor. This work has enabled us to test our synthetic and biological met...
Article
Full-text available
Insulin and insulin-like growth factors I and II are closely related protein hormones. Their distinct evolution has resulted in different yet overlapping biological functions, with insulin becoming a key regulator of metabolism, while IGF-I/II are major growth factors. Insulin and IGFs cross-bind with different affinities to closely related insulin...
Article
Insulin, insulin-like growth factors 1 and 2 (IGF-1/-2), and their receptors (IR, IGF-1R), are the key elements of complex hormonal system that is essential for development and functioning of humans. The C and D domains of IGFs (absent in insulin) are likely playing important roles in the differential binding of IGF-1/-2 to IGF-1R, the isoforms of...
Article
Insulin, insulin-like growth factors 1 and 2 (IGF-1 and -2, respectively), and their receptors (IR and IGF-1R) are the key elements of a complex hormonal system that is essential for the development and functioning of humans. The C and D domains of IGFs (absent in insulin) likely play important roles in the differential binding of IGF-1 and -2 to I...
Article
Full-text available
Insulin is a key hormone of human metabolism with major therapeutic importance for both types of diabetes. New insulin analogues with more physiological profiles and better glycemic control are needed, especially analogues that preferentially bind to the metabolic B-isoform of insulin receptor (IR-B). Here, we aimed to stabilize and modulate the re...
Article
Full-text available
The insulin gene mutation c.137G>A (R46Q), which changes an arginine at the B22 position of the mature hormone to glutamine, causes the monogenic diabetes variant maturity-onset diabetes of the young (MODY). In MODY patients, this mutation is heterozygous, and both mutant and wild-type (WT) human insulin are produced simultaneously. However, the pa...
Article
Full-text available
The structural characterization of the insulin–insulin receptor (IR) interaction still lacks the conformation of the crucial B21–B30 insulin region, which must be different from that in its storage forms to ensure effective receptor binding. Here, it is shown that insulin analogues modified by natural amino acids at the TyrB26 site can represent an...
Article
The N-terminus of the B-chain of insulin may adopt two alternative conformations designated as the T- and R-states. Despite the recent structural insight into insulin:insulin receptor (IR) complexes, the physiological relevance of the T/R transition is still unclear. Hence this study focused on the rational design, synthesis and characterization of...
Article
Full-text available
Until recently, despite decades of intensive research, the exact mode of interaction of insulin with its membrane receptor remained elusive. However, recent advances achieved with insulin analogues and crystallographic studies with the insulin receptor provided a first insight into the interaction of insulin with its primary binding site on the ins...
Article
Full-text available
Despite the recent first insight into the insulin-insulin receptor complex, the structural role of the C-terminus of the B-chain of insulin in this assembly remains unresolved. Previous studies have suggested that this part of insulin must rearrange to reveal amino acids crucial for interaction with the receptor. The role of the invariant PheB24, o...
Article
Full-text available
Insulin receptor signalling has a central role in mammalian biology, regulating cellular metabolism, growth, division, differentiation and survival. Insulin resistance contributes to the pathogenesis of type 2 diabetes mellitus and the onset of Alzheimer's disease; aberrant signalling occurs in diverse cancers, exacerbated by cross-talk with the ho...
Article
Full-text available
The CART (cocaine- and amphetamine-regulated transcript) peptide is an anorexigenic neuropeptide that acts in the hypothalamus. The receptor and the mechanism of action of this peptide are still unknown. In our previous study, we showed that the CART peptide binds specifically to PC12 rat pheochromocytoma cells in both the native and differentiated...
Article
Full-text available
Apart from its role in insulin receptor (IR) activation, the C terminus of the B-chain of insulin is also responsible for the formation of insulin dimers. The dimerization of insulin plays an important role in the endogenous delivery of the hormone and in the administration of insulin to patients. Here, we investigated insulin analogues with select...
Article
Full-text available
Apart from its role in insulin receptor (IR) activation, the C terminus of the B-chain of insulin is also responsible for the formation of insulin dimers. The dimerization of insulin plays an important role in the endogenous delivery of the hormone and in the administration of insulin to patients. Here, we investigated insulin analogues with select...
Article
Full-text available
Insulin is a key protein hormone that regulates blood glucose levels and, thus, has widespread impact on lipid and protein metabolism. Insulin action is manifested through binding of its monomeric form to the Insulin Receptor (IR). At present, however, our knowledge about the structural behavior of insulin is based upon inactive, multimeric, and st...
Article
In this study, we prepared several shortened and full-length insulin analogues with substitutions at position B26. We compared the binding affinities of the analogues for rat adipose membranes with their ability to lower the plasma glucose level in nondiabetic Wistar rats in vivo after subcutaneous administration, and also with their ability to sti...
Article
Chromatographic behavior of a series of N-methylated tetra and octapeptides on a reversed-phase sorbent was studied considering the information obtained on these compounds by NMR spectroscopy. The modified tetrapeptides were derived from GFFY-NH2, GFFF-NH2 and GFFH-NH2 primary structures by N-methylation at various peptide bond positions. Similarly...
Article
In this paper, we present the detailed synthetic protocol and characterization of Fmoc-Lys(Pac)-OH, its use for the preparation of octapeptides H-Gly-Phe-Tyr-N-MePhe-Thr-Lys(Pac)-Pro-Thr-OH and H-Gly-Phe-Phe-His-Thr-Pro-Lys(Pac)-Thr-OH by solid-phase synthesis, trypsin-catalyzed condensation of these octapeptides with desoctapeptide(B23-B30)-insuli...
Article
A direct and convenient spectrophotometric assay has been developed for methionine aminopeptidases (MetAPs). The method employs the hydrolysis of a substrate that is a methionyl analogue of p-nitroaniline (L-Met-p-NA), which releases the chromogenic product p-nitroaniline. This chromogenic product can be monitored continuously using a UV-Vis spectr...
Article
Full-text available
Despentapeptide (des-B26-B30) insulin (DPI), an active modified insulin, has been crystallized in the presence of 20% acetic acid pH 2. A crystal structure analysis to 1.8 A spacing (space group I222) revealed that the DPI molecule, which is unable to make beta-strand interactions for physiological dimer formation and is apparently monomeric in sol...
Article
Insulin was isolated and used as a therapeutic agent for the first time more than 80 years ago. In subsequent decades, the structure of insulin was fully resolved and several insulin analogues were introduced in clinical practice. Important discoveries - were made concerning the mechanism of action of insulin, its interaction with receptors, and in...
Article
The origins of differentiation of insulin from insulin-like growth factor I (IGF-I) are still unknown. To address the problem of a structural and biological switch from the mostly metabolic hormonal activity of insulin to the predominant growth factor activities of IGF-I, an insulin analogue with IGF-I-like structural features has been synthesized....
Article
Capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) were used for the analysis of new synthetic derivatives of hypophysis neurohormones--vasopressin and oxytocin, and pancreatic hormone--human insulin (HI) and its octapeptide fragment, derivatized by fluorescent probe, 4-chloro-7-nitrobenzo[1,2,5]oxadiazol (NBD)....
Article
The preparation and characterization of two novel LysB29 selectively labelled fluorescent derivatives of human insulin are described. Two probes were chosen: 4-chloro-7-nitrobenz-2-oxa-1,3-diazole (NBD) and 7-methoxycoumarin-4-acetic acid (MCA), which have a relatively small, compact structure and are able to react with amino groups to form highly...
Article
The role of three highly conserved insulin residues PheB24, PheB25, and TyrB26 was studied to better understand the subtleties of the structure-function relationship between insulin and its receptor. Ten shortened insulin analogues with modifications in the beta-strand of the B-chain were synthesized by trypsin-catalyzed coupling of des-octapeptide...
Article
An ever-icreasing number of patients who suffer from diabetes mellitus is a worldwide problem. Therefore, an extensive study of insulin and its receptor is of high priority of many recent studies. Particularly, a detailed knowledge of the molecular structure and conformational changes of insulin along the metabolic path is very important. The paper...