Topic 4  Immunoglobulins: Structure and Function

Introduction

One of the major functions of the immune system is the production of soluble proteins that circulate freely and exhibit properties that contribute specifically to immunity and protection against foreign material.  These soluble proteins are the antibodies, and they belong to the class of proteins called globulins.  Most of this section focuses on how the primary, secondary and tertiary structure of immunoglobulins contribute to both their  specificity  and their effector functions.  The antibody molecule has evolved to perform two distinct functions- antigen recognition and antigen elimination. Antibody molecules can interact with a virtually unlimited number of antigens, yet they destroy or eliminate antigens by a small number of effector mechanisms. We will examine each major class of immunoglobulin, describing its structure and key features.  Finally we’ll spend a little time describing the tumors of plasma cells called myeloma and engineering of monoclonal antibodies.

Objectives

On completion of this section and the required readings, you should be able to:

n  distinguish between the overall structure and the fine structure of immunoglobulins;

n  describe the variable and constant regions of immunoglobulin’s light and heavy chains;

n  explain the structural organization of the variable regions of an immunoglobulins light and heavy chains;

n  differentiate between hypervariable regions and complementarity determining regions;

n  contrast monoclonal antibody production vs polyclonal antibody production;

n  discuss the differences in the biological effector functions of the different classes of immunoglobulins;

n  draw a diagram of the procedure for monoclonal antibody production;

n  construct a table comparing the biological characteristics of the five classes of immunoglobulins;

n  construct a table comparing the biochemical and biophysical characteristics of the five classes of immunoglobulins.

Required Reading

Please refer to the textbook key for specific readings for this section.

P Key Terms

•    allotypes, •    allotypic determinants, •    Am determinants, •    Gm determinants, •    Km determinants, •    Bence Jones proteins •    immunoglobulin, classes, •    antibody subclasses, •    constant region (C), •    Fab fragment, •    Fab’ fragment, •    F(ab’)2 fragment, •    Fc fragment, •    Framework residues (FR), •    heavy chain, •    hinge regions, •    chimeric antibodies, •    hybridoma

•    hypervariable regions (HVRs), •    complementarity determining regions (CDRs) •    Hypoxantinine •    Aminopterin •    Thymidine medium (HAT) •    idiotypes, •    idiotypic determinants •    immunoglobulin domains •    immunoglobulins •    isotype, •    isotypic determinants •    joining chain (J), •    Light chain •    monoclonal antibodies, •    multiple myeloma •    Secretory IgA •    Segmental flexibility, •    variable regions (V)

P Key Concepts

n  Immunoglobulins of all classes have a fundamental four chain structure, consisting of two identical light (L) and two identical heavy chains, which are held together by disulfide bonds.

n  Each of the domains in the immunoglobulin molecule has a characteristic tertiary structure called the immunoglobulin fold.  Constant (C) region domains are similar within each class of immunoglobulin molecules.  The N-terminal domains of both heavy and light chains are the variable (V) regions, which together make up the combining site of the antibody.

n  The classes of immunoglobulin molecules differ by virtue of the constant regions formed from their H chains, which are responsible for the different biologic functions carried out by each class.

n  Unlike polyclonal antibodies, monoclonal antibody is a homogenous preparation specific for a single epitope on a complex antigen.

DID YOU KNOW?

Immunoglobulins function as antibodies, the antigen - binding proteins present on the B-cells membrane and secreted by plasma cells.  Secreted antibodies circulate in the blood and serve as the effectors of humoral immunity by searching out and neutralizing or eliminating antigens.  All immunoglobulins share certain structural features, bind to antigen, and participate in a limited number of effector functions.  The knowledge of the structure-function relations in immunoglobulins helped develop new treatments of diseases caused by the failing immune system. However in spite of a tremendous amount of research and multiple approaches used by researchers,  solving one of the most difficult problems of modern medicine - developing a vaccine for cancer, proves to be a great challenge.  Some hope however seems to appear on the horizon:

Healing Cancer - Vaccines that prod the body to cure itself are finally being readied for market.

If all goes as expected, the first vaccine against cancer will be approved for sale before the end of September 1998.  The vaccine will neither prevent cancer nor cure it, and it would first be sold in Canada, not the U.S.  It will be a significant event nonetheless, because it will demonstrate that a long-held dream - of attacking cancer by guile from within, rather than assaulting the body by brute force from without - is beginning to come true. 

More than half a dozen large-scale tests of cancer vaccines are under way in clinics around the world.  Most aim at the same malignancy as this first drug: melanoma, a fast-spreading skin cancer that strikes about one person in 100.

Ribi ImmunoChem Research, a biotech firm in Hamilton, Mont., was simply the first to file for market approval.  European regulators are also evaluating the company’s clinical results, and Ribi plans to put its new medicine, Melacine, before U.S. Food and Drug Administration reviews later this year. 

What those experts will see is evidence that nearly all the 70 terminal patients injected with this cocktail of ripped-up tumor cells and bits of bacteria that cause tuberculosis felt significantly fewer ill effects than the 70 given standard chemotherapy. The vaccine made patients’ lives easier but not longer.  At least not on average; the lucky few who responded well to the vaccine did survive longer than those who responded well to conventional drugs.

There is good reason to hope that other, more sophisticated vaccines still in clinical trials will improve on those modest gains, in two ways.  They may contain more potent adjuvants, additives such as the bacterial fragments in Melacine that awaken the body’s immune system to the fact that the cancer doesn’t belong there.  And they may use more effective antigens, fragments of tumor cells that train antibodies and killer T cells to recognize cancer when they see it.

In March, for example, Steven A. Rosenberg and his colleagues at the National Cancer Institute reported good news about a vaccine they have made from a particular protein fragment and interleukin-2, a chemical secreted by T cells when they stumble on foreign bodies.  Of the 31 patients with widespread melanoma who were immunized with the new medicine, 13 saw their tumors shrink by more than half.

Rosenberg’s group went to great efforts just to identify just the right section of protein to use in this vaccine, but a shotgun technique may also work against some cancers.  Michael G. Hanna, chairman of Intracel in Rockville, MD., announced in July that a decade-long test of its ONCOVaxCL vaccine for colon cancer had succeeded.  Intracel borrowed parts of tumors removed from patients’ colons, digested them with enzymes and then injected each patient with his or her own tumor cells, along with a bacterial adjuvant.  In people suffering from stage II colon cancer, the vaccine appeared to cut the rate at which the disease resurged by 61 percent over about five years, when compared with patients treated by surgery alone.  Intracel is planning to file for FDA approval of its drug late this year.

Other large scale vaccine trails are just getting started.   Progenics Pharmaceuticals in Tarrytown, N.Y., had by July enrolled more than half of 800 skin cancer patients it wants for its study.  They will test a concoction of a carbohydrate antigen and an adjuvant derived from the bark of the South American soap tree, says Robert J. Israel, the company’s chief scientist.

ImClone Systems in New York City is gearing up for a trial of similar size to see whether its vaccine will prevent the recurrence of small-cell lung cancer.  “Virtually all patients with this disease relapse after their initial treatment,” says Harlan W. Waksal, ImClone’s chief operating officer.  “The disease usually comes back within a year - and with a vengeance.  We hope to stop that.”  ImClone’s antigen might seem like an unlikely champion, constructed as it was by making an antibody to an antibody of sugar-fat compound on cancer cells.  But in small-scale trials, Waksal reports, about 40 percent of people given the vaccine survived five years, despite odds predicting that fewer than 5 percent of them would hang in that long. 

The John Wayne Cancer Institute in Santa Monica, California,  is coordinating an even larger and longer study, to span five years and eight nations and to include 1,100 people whose melanoma has spread into their lymphatic system.  The subjects will be given either irradiated melanoma tumor cells or interferon alfa-2b, a drug that forces tumors to display the antigens that make them susceptible to attack.  (The two drugs need not be exclusive; Ribi is running a clinical trial to see whether they work well together.)

The largest trial of cancer vaccine so far, however, is unfortunately an unscientific one.  In the past year, reportedly upward of 50,000 people in China have been injected with kang lai te, an extract from seeds of the herb Job’s tears (Coix lacryma-jobi) that the government has endorsed as treatment for cancers of the lung, liver and stomach. 

It is too early to say whether science can coax and coach the human body to defend itself successfully against itself.  But at the very least, medicine now seems poised to offer a more palatable exit strategy than poison, radiation or the blade.

W. Wayt Gibbs Scientific American, September 1998, pp 40-41

Review Questions

1. Textbook Study Questions

Review questions at the end of the Chapter 5.  The answers with explanations are available at the end of the textbook.

2. Multiple Choice Questions

1.  The specificity of an antibody is due to

A) The constant portions of the H and L chains.

B)  The variable portions of the H and L chains.

C)  Its valence.

D) The L chains.

E)  The H chains.

 

 

 

2.  Which statement is INCORRECT?

A) The Fc region attaches to a host cell.

B)  The constant region of a heavy chain is the same for all antibodies.

C)  The variable region of a heavy chain binds with antigen.

D) The variable region of a light chain binds with antigen

E)  None of these.

3.  Reaction of antigen with IgE antibodies attached to mast cells causes

A) Complement fixation.

B)  Agglutination.

C)  Lysis of the cells.

D) Release of chemical mediators.

E)  None of these.

4.  Antibodies for serological testing can be obtained from all of the following except

A) Vaccinated humans.

B)  None of these.

C)  Hybridoma clones.

D) Vaccinated animals

E)  Viral cultures.

5.  Serum IgM molecules usually have

A) 10 light chains

B)  10 heavy chains

C)  a pentameric structure

D) a J chain

E)  all of the above

3. Definition/Short Answer Questions

1.  The analysis of IgG molecules after hydrolysis by pepsin and papain led to similar yet different results. Explain.

2.  What is the difference between and immunoglobulin and a myeloma protein?

3.  Why were myeloma proteins and Bence Jones proteins critical to the early studies on antibody structure? What has largely replaced them?

4.  Differentiate among the following: complementarity determining regions, hyper-variable regions and framework regions.

5.  Explain the statements:

“Antibodies can be antigens” and

“Antibodies can be used to characterize antibodies.”

6.  Discuss the terms immunoglobulin isotypes, immunoglobulin allotypes and immunoglobulin idiotypes and give examples of each.

7.  Compare conventional antibody production with monoclonal antibody production and recombinant antibody production.

8.  What are antibody domains?

9.  Why can’t light chains be used to classify antibodies?