Biological Chemistry - Berezov, T. T., Korovkin, B. F. 1998

Hormones
Pancreatic Hormones
Insulin

The Pancreas is a gland with mixed secretion. Its exocrine function involves the synthesis of A number of key digestive Enzymes—specifically amylase, lipase, Trypsin, Chymotrypsin, carboxypeptidase, and others—which enter the intestine via pancreatic juice. The endocrine function is performed by the pancreatic islets (islets of Langerhans), as discovered in 1902 by L.V. Sobolev. These islets consist of various Cell types that produce Hormones with generally opposing effects. For instance, a- (or A-) Cells produce Glucagon, ß- (or B-) cells synthesize Insulin, d- (or D-) cells secrete Somatostatin, and F-cells produce the Pancreatic Polypeptide, which is less well understood. Insulin and glucagon will be examined below as hormones of exceptional importance for the vital activity of the Organism*.

* Available data regarding somatostatin as a hypothalamic hormone (see above) indicate that the pancreas and a number of intestinal cells secrete their own somatostatin, which exerts an inhibitory effect on the secretion of insulin and glucagon by the a- and ß-Cells of the islets of Langerhans.

Insulin, named after the pancreatic islets (from Latin insula meaning island), was the first protein whose Primary Structure was elucidated in 1954 by F. Sanger (see Chapter 1). Pure insulin was obtained in 1922 following its discovery in extracts of pancreatic islets by F. Banting and C. Best. The insulin molecule, containing 51 amino acid residues, consists of two polypeptide chains interconnected at two points by disulfide bridges. The structure of insulin and its precursor, proinsulin, is shown in Chapter 1 (see Fig. 1.14). Currently, it is standard to designate the 21-membered peptide as the A-chain of insulin and the peptide containing 30 amino acid residues as the B-chain. Furthermore, the Chemical synthesis of insulin has been accomplished in many laboratories. Porcine insulin is structurally closest to human insulin, differing only in that Threonine at position 30 of the B-chain is replaced by Alanine.

There are no significant differences in the Amino Acid Sequence of insulin among various animals. Insulins differ in the Amino Acid Composition of the A-chain at positions 8–10.

According to current concepts, The Biosynthesis of insulin in the ß-cells of the pancreatic islets proceeds from its precursor, proinsulin, which was first isolated by D. Steiner in 1966. Today, not only has the Introduction/19.html">Primary structure of proinsulin been elucidated, but its chemical synthesis has also been achieved (see Fig. 1.14). Proinsulin is represented by a single polypeptide chain containing 84 amino acid residues and lacks biological, i.e., hormonal, activity. The microsomal fraction of the pancreatic islet ß-cells is considered the site of proinsulin synthesis; The conversion of inactive proinsulin into active insulin (the most crucial part of synthesis) occurs during The transport of proinsulin from Ribosomes to secretory granules via partial proteolysis (Cleavage from the C-terminus of the polypeptide chain of a peptide containing 33 amino acid residues, known as the connecting peptide, or C-peptide). The length and primary STRUCTURE OF THE C-peptide exhibit greater Variability across animal species than the sequences of the insulin A and B chains. It has been established that the initial precursor of insulin is preproinsulin, which contains, In addition to proinsulin, a so-called leader or signal sequence at its N-terminus consisting of 23 amino acid residues. During The formation of the proinsulin molecule, this signal peptide is cleaved by a specialized peptidase. Subsequently, the proinsulin molecule also undergoes partial proteolysis, and under the action of a trypsin-like proteinase, two basic Amino Acids are cleaved from the N- and C-termini of the C-peptide—specifically the Arg—Arg and Lys—Arg dipeptides, respectively (see Fig. 1.14). However, The Nature of the enzymes and the subtle mechanisms of this vital biological process—the Formation of the active insulin molecule—have not yet been completely elucidated.

Insulin synthesized from proinsulin can exist in several forms that differ in their biological, immunological, and physicochemical properties. Two forms of insulin are distinguished: 1) free insulin, which interacts with Antibodies raised against crystalline insulin and stimulates glucose uptake by Muscle and adipose Tissues; and 2) bound insulin, which does not react with antibodies and is active solely with respect to adipose tissue. The existence of the bound insulin form has now been proven, and its localization within Blood serum protein fractions—specifically in the transferrin and a-globulin regions—has been established. The Molecular Weight of bound insulin ranges from 60,000 to 100,000. In addition, there is the so-called form A of insulin, which differs from the previous two in a number of physicochemical and biological properties, occupies an intermediate position, and appears in response to the rapid, urgent demand of the organism for insulin.

Blood glucose concentration plays a dominant role in the physiological Regulation of Insulin synthesis. Thus, an increase in blood glucose content causes an elevation of insulin secretion in the pancreatic islets, whereas a decrease leads to a suppression of insulin secretion. This feedback control phenomenon is regarded as one of the most critical mechanisms for regulating blood glucose levels. Furthermore, Insulin secretion is influenced by electrolytes (especially Calcium Ions), amino acids, glucagon, and secretin. Evidence supporting The Role of the cyclase system in insulin secretion is also presented. It is hypothesized that glucose acts as a signal to activate adenylate cyclase, and the cAMP generated in this system serves as the signal for insulin secretion.

Inadequate secretion (or more precisely, insufficient synthesis) of insulin leads to a specific disorder: Diabetes Mellitus (see Chapter 10). In addition to clinical symptoms (polyuria, polydipsia, and polyphagia), diabetes mellitus is characterized by a series of specific metabolic disturbances. For example, patients develop hyperglycemia (elevated blood glucose levels) and glucosuria (The excretion of glucose in urine, where it is normally absent). Metabolic Disorders also include enhanced Glycogen breakdown in The Liver and Muscles, slowed Biosynthesis of Proteins and Lipids, a reduced rate of glucose oxidation in tissues, The Development of a negative nitrogen balance, and increased blood levels of Cholesterol and other lipids. In diabetes, fat mobilization from fat depots, carbohydrate synthesis from amino acids (Gluconeogenesis), and the excessive synthesis of Ketone Bodies (ketonuria) are intensified. Following insulin administration to patients, all of these abnormalities generally disappear; however, the hormone's action is time-limited, making continuous administration necessary. The clinical symptoms and metabolic disturbances in diabetes mellitus cannot be explained solely by the lack of insulin synthesis. Evidence has been obtained that In the second form of diabetes mellitus—the so-called insulin-resistant form—molecular defects also occur, specifically aberrations in insulin structure or in the enzymatic conversion of proinsulin to insulin. The development of this form of diabetes is frequently rooted in the loss of the target cells' receptors' ability to bind to the structurally aberrant insulin molecule, or in the synthesis of a mutant receptor (see below).

In experimental animals, insulin administration induces hypoglycemia (a drop in blood glucose levels), an increase in muscle glycogen stores, the enhancement of anabolic processes, and an elevated rate of glucose utilization in tissues. Additionally, insulin exerts an indirect effect on Water and Mineral METABOLISM.

The MECHANISM OF ACTION of insulin has not been fully deciphered, despite a vast amount of empirical data demonstrating a close and direct relationship between insulin and metabolic processes in the organism. According to the "unitary" theory, all effects of insulin are mediated through its influence on glucose metabolism via the enzyme hexokinase. New experimental data indicate that the enhancement and stimulation by insulin of processes such as ion and Amino Acid Transport, Translation and Protein Synthesis, Gene Expression, and others are independent. This has served as the basis for the hypothesis regarding multiple mechanisms of insulin action.

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Fig. 8.1. Insulin Receptor (schematic diagram).

Two a-chains on the outer surface of The Cell membrane and two transmembrane ß-chains. Insulin binding to the a-chains triggers autophosphorylation of Tyrosine residues within the ß-chains; the active tyrosine kinase domain subsequently participates in the phosphorylation of inactive target proteins in the Cytosol.

At present, the membrane localization of the primary action of nearly all protein hormones, including insulin, appears most probable. Evidence has been obtained for the existence of a specific insulin receptor on the outer Plasma Membrane of nearly all cells in the organism, as well as for the formation of an insulin-receptor complex. The receptor is synthesized as a precursor polypeptide (1,382 amino acid residues, MW 190,000), which is subsequently cleaved into a- and ß-subunits—forming a heterodimer (with the formula a₂—ß₂) linked by Disulfide Bonds. It has been found that while the a-subunits (MW 135,000) are located almost entirely on the outer surface of the biomembrane, performing the function of binding cellular insulin, the ß-subunits (MW 95,000) represent a transmembrane protein that carries out signal Transduction (Fig. 8.1). The concentration of insulin receptors On the surface reaches 20,000 per cell, and their half-life is 7–12 hours.

The most fascinating property of the insulin receptor, distinguishing it from all other protein and peptide Hormone Receptors, is its ability to undergo autophosphorylation—meaning the receptor itself possesses protein kinase (tyrosine kinase) activity. Upon binding of insulin to the a-chains of the receptor, the tyrosine kinase activity of the ß-chains is activated through the phosphorylation of their tyrosine residues. In turn, the active tyrosine kinase of the ß-chains triggers a cascade of phosphorylation-dephosphorylation of protein Kinases, specifically membrane- or cytosol-bound Serine or threonine kinases (i.e., protein kinases and target proteins in which phosphorylation is mediated via the OH groups of Serine and threonine). Consequently, changes in cellular activity occur, notably the Activation and inhibition of enzymes, glucose transport, the synthesis of nucleic acid and protein polymers, and so forth*.

It must be emphasized, however, that the subtle MOLECULAR MECHANISMS OF the signal transduction pathways from the insulin-receptor complex to numerous intracellular processes have not yet been elucidated. It is quite plausible that a number of intracellular second messengers participate in such processes, particularly Cyclic NUCLEOTIDES, phosphatidylinositol derivatives, and others. Furthermore, one cannot rule out the possibility of the existence of an intracellular mediator or modulator of insulin action (a specialized intracellular receptor) that controls gene Transcription and, accordingly, mRNA synthesis. It is hypothesized that insulin's action and its involvement in regulating gene expression or the transcription of specific mRNAs may explain its role in such fundamental life processes as Embryogenesis and the differentiation of cells in higher organisms.