ADVANCES IN THE STUDY
OF GENETIC DISORDERS

Edited by Kenji Ikehara Advances in the Study of Genetic Disorders
Edited by Kenji

Ikehara Published by InTech
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p. cm.
ISBN 978-953-307-305-7 Contents

Preface IX
Part 1 Background of Genetic Disorder 1
Chapter 1 Origin of the Genetic Code and Genetic Disorder 3
Kenji Ikehara
Chapter 2 Inbreeding and Genetic Disorder 21
Gonzalo Alvarez, Celsa Quinteiro and Francisco C. Ceballos
Chapter 3 Cytogenetic Techniques in Diagnosing Genetic Disorders 45
Kannan Thirumulu Ponnuraj
Chapter 4 Functional Interpretation of Omics Data by Profiling Genes
and Diseases Using MeSH–Controlled Vocabulary 65
Takeru Nakazato, Hidemasa Bono and Toshihisa Takagi
Chapter 5 Targeted Metabolomics for Clinical Biomarker Discovery in
Multifactorial Diseases 81
Ulrika Lundin, Robert Modre-Osprian and Klaus M. Weinberger
Part 2 Unifactorial or Unigenetic Disorder 99
Chapter 6 Thalassemia Syndrome 101
Tangvarasittichai Surapon
Chapter 7 Genomic Study in β-Thalassemia 149

Part 3 Multifactorial or Polygenic Disorder 319
Chapter 16 Peroxisomal Biogenesis:
Genetic Disorders Reveal the Mechanisms 321
Manuel J. Santos and Alfonso González
Chapter 17 Repair of Impaired Host Peroxisomal Properties Cropped Up
Due to Visceral Leishmaniasis May Lead to Overcome
Peroxisome Related Genetic Disorder Which May Develop
Later After Treatment 333
Salil C. Datta, Shreedhara Gupta and Bikramjit Raychaudhury
Chapter 18 Genetic Basis of Inherited Bone Marrow
Failure Syndromes 357
Yigal Dror
Chapter 19 Bernard Soulier Syndrome: A Genetic Bleeding Disorder 393
Basma Hadjkacem, Jalel Gargouri and Ali Gargouri
Contents VII

Chapter 20 Prader–Willi Syndrome, from Molecular Testing and Clinical
Study to Diagnostic Protocols 409
Maria Puiu and Natalia Cucu
Chapter 21 Turner Syndrome and Sex Chromosomal Mosaicism 431
Eduardo Pásaro Méndez and Rosa Mª Fernández García
Chapter 22 Microstomia: A Rare but Serious Oral Manifestation of
Inherited Disorders 449
Aydin Gulses

human beings from many kinds of diseases caused by the infection of viruses and
bacteria, resulting in extending the life span of human beings. Currently, many
Japanese people can live until between 90 and 100 years old. For example, the average
life spans of females and males living in Japan had reached 86.4 and 79.6 years old by
2009, respectively, while the comparative figures in 1950 were about only 62 and 58
years old, respectively.
X Preface

It is reported that the highest cause of Japanese deaths is malignant tumour or cancer.
Cancers induced by genetic defects leading to deviation from the normal control of cell
division can be regarded as a kind of genetic disorder. The genetic defects may occur
in all organs, such as the kidney, the spleen, the stomach, the lung and the intestine
etc. In addition, it is quite difficult to cure these cancers by the usual treatments such
as administration of medicines (except for removal of malignant tumours by surgical
operations) because at the present time it is impossible to site-specifically replace the
substituted bases to the original/normal bases. This is the reason why cancers are at
the top of the Japanese death causes although human beings are released from many
kinds of infectious diseases.
Many genetic disorders are caused by base substitutions on double-stranded DNA, as
with cancers. Although the mutated bases must be replaced with the original/normal
bases in order to completely cure the disorders, it is quite difficult to achieve this
purpose at the present time, again, as with the case of cancers as described above.
Thus, genetic disorders remain diseases which are difficult to cure. In addition,
mutations causing genetic disorders may occur in any cells carrying genetic elements
or DNA and at anytime. Therefore, the organisms living on earth have been exposed
to danger-generating base substitutions without exception, and genetic disorders may
be induced in any organs because human beings are multi-cellular organisms.
There are two big problems with genetic disorders. One is that it is quite difficult to
cure them, as described above. However, in addition to the knowledge about such
mechanisms as DNA replication, transcription and the translation of genetic

Watson and Crick in 1956. Therefore, nowadays it is possible to understand the
reasons why genetic disorders are caused. It is probable that the knowledge of genetic
disorders described in this book will lead to the discovery of an epoch of new medical
treatment and relieve human beings from the genetic disorders of the future, because
human beings had overcome many difficulties already (such as infectious diseases
through the discovery of new medical treatment using vaccines for protection against
infection form viruses and of special medicines known as antibiotics for curing
diseases caused by the infection of micro-organisms). As such, I have a presentiment
that a new age is now dawning with respect to the overcoming of genetic disorders.
The dawn may set in suddenly upon a big discovery for a new medical treatment -
which will be achieved by one genius in the future - because such kinds of big
discoveries have always been carried out suddenly by geniuses, such as Jenner and
Fleming. I hope that the descriptions in this book will contribute to such a discovery,
of a new medical treatment for genetic disorders.

Kenji Ikehara
The Open University of Japan, Nara Study Centre,
International Institute for Advanced Studies of Japan
Japan
Part 1
Background of Genetic Disorder

1
Origin of the Genetic Code
and Genetic Disorder
Kenji Ikehara
The Open University of Japan, Nara Study Center

on, which affect various functions for gene expression leading to synthesis of lower or
higher amounts of proteins than normal level, resulting in many kinds of genetic diseases
(Figure 1 (A)).

Advances in the Study of Genetic Disorders
4
(A)

(B)

Fig. 1. (A) Possible mutation sites, which may affect various functions for gene expression
and catalytic functions of proteins. Dark and white horizontal bars indicate exons encoding
amino acid sequences of a protein and introns without genetic information for protein
synthesis, respectively. Capital letters, P and T, mean a promoter for transcription initiation
and a terminator required for termination of mRNA synthesis, respectively. Thick upward
open and closed arrows and thin downward arrows indicate insertion and deletion of DNA
sequences, and one-base substitutions, respectively. (B) Amino acid replacement observed
in a classical and well-known genetic disorder, sickle cell anemia. Red letters indicate
replacements of amino acid and base of the genetic mRNA sequence

Genetic Disorder Inheritance Gene
Hailey-Hailey Disease Autosomal dominant ATP2C1
Adenosine deaminase deficiency Autosomal recessive ADA
Thalassemia globins
Alstrom Syndrome ALMS1
Tangier Disease ABCA1
Phenylketourea PAH
Galactosemia GALT
Aicardi-Goutieres syndrome X-link dominant RNAses
Bernard-Soulier syndrome GPIs

such as about 25 years in the case of human, have occurred at a comparatively low
frequency. On the other hand, amino acids of microbial proteins have been substituted at
a high frequency without largely affecting protein functions. That is because evolution
rate of microbial proteins is quite large due to the enormously large cell number and a
quite short division time, such as about 20-30 minutes in the case of Escherichia coli.
Therefore, it would be suitable to compare an amino acid sequence of a microbial protein
with the homologous amino acid sequence in order to investigate amino acid
substitutions occurring without largely affecting the protein function in a wide range as
shown in Figure 2. Fig. 2. Alignment of two amino acid sequences of small homologous single-stranded DNA
binding proteins, from Aquifex aeolicus (147 amino acids) and Carboxydothermus
hydrogenoformans (142 amino acids). Red bold and black letters indicate substituted and
conserved amino acids between the two amino acid sequences, respectively. Hyphen (-)
means amino acid position deleted from one amino acid sequence. Homology percent
between the two single-stranded DNA binding proteins, which were obtained from
GeneBank at is 38%

Advances in the Study of Genetic Disorders
6
A C D E F G H I K L M N P Q R S T V W Y
A 0,0 4,0 6,0 0,0 1,2 2,0 2,0 1,0 2,0 2,0 4,0 1,0 2,0 3,1 6,0 2,0 4,1 0,0 3,0
C 0,0 0,0 0,0 0,0 0,0 0,0 1,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0
D 0,0 1,0 5,1 1,0 1,0 0,0 0,0 4,0 1,0 2,0 2,0 0,0 3,0 0,0 2,0 2,1 0,0 0,0 0,0
E 1,0 0,0 1,5 1,1 0,1 0,0 1,1 5,0 0,1 1,0 1,1 1,1 3,0 3,2 2,3 2,1 1,0 0,0 2,0
F 0,0 0,0 0,0 0,0 0,0 0,0 2,3 0,0 1,1 0,0 0,0 0,0 1,0 1,1 0,0 0,0 1,0 0,0 5,0
G 1,0 0,0 1,0 1,0 0,0 0,0 0,0 5,0 0,0 0,0 3,1 0,0 2,1 1,1 2,0 1,0 0,0 0,0 1,0
H 1,0 0,0 1,1 1,0 0,0 1,0 0,0 0,0 0,0 0,0 2,0 0,0 0,0 0,0 0,0 0,0 0,0 0,0 1,0
I 0,0 0,0 0,0 1,0 0,0 0,0 0,0 0,0 3,3 1,0 0,0 0,1 0,0 0,0 0,0 0,0 7,3 0,0 1,0

Origin of the Genetic Code and Genetic Disorder
7
As seen in Figure 2, many amino acid substitutions are observed between two homologous
single-stranded DNA binding proteins. The amino acid substitutions caused by base
substitutions at the first codon position were observed more than those caused by base
substitutions at the second codon position (see the Table given in Figure 3). Similar results
were obtained from amino acid substitutions between two large homologous stringent
response proteins, Streptomyces coelicolor RelA and Staphylococcus aureus RelA (Figure 3). It
can be interpreted as that amino acids with similar chemical and physical properties are
arranged in the same column in the genetic code table at a comparably high probability
(Table 2 (A), (B), (C) and (D)).
The universal genetic code is redundant and has a highly non-random structure. Typically,
when nucleotide at the third codon position differs from the corresponding one, both
codons encode the same amino acids at a high probability, due to the degeneracy of the
genetic code at the third codon position. In addition, codons, of which nucleotide at the first
codon position differs from each other, usually encode amino acids with different but rather
similar chemical/physical properties.

(A) (B)
Hydropathy
α-Helix
U C A G U C A G
Phe Ser Tyr Cys U Phe Ser Tyr Cys U
U Phe Ser Tyr Cys C U Phe Ser Tyr Cys C
Leu Ser Term Term A Leu Ser Term Term A
Leu Ser Term Trp G Leu Ser Term Trp G
Leu Pro His Arg U Leu Pro His Arg U
C Leu Pro His Arg C C Leu Pro His Arg C
Leu Pro Gln Arg A Leu Pro Gln Arg A
Leu Pro Gln Arg G Leu Pro Gln Arg G

(C) (D)
β-Sheet Turn/Coil
U C A G U C A G
Phe Ser Tyr Cys U Phe Ser Tyr Cys U
U Phe Ser Tyr Cys C U Phe Ser Tyr Cys C
Leu Ser Term Term A Leu Ser Term Term A
Leu Ser Term Trp G Leu Ser Term Trp G
Leu Pro His Arg U Leu Pro His Arg U
C Leu Pro His Arg C C Leu Pro His Arg C
Leu Pro Gln Arg A Leu Pro Gln Arg A
Leu Pro Gln Arg G Leu Pro Gln Arg G
Ile Thr Asn Ser U Ile Thr Asn Ser U
A Ile Thr Asn Ser C A Ile Thr Asn Ser C
Ile Thr Lys Arg A Ile Thr Lys Arg A
Met Thr Lys Arg G Met Thr Lys Arg G
Val Ala Asp Gly U Val Ala Asp Gly U
G Val Ala Asp Gly C G Val Ala Asp Gly C
Val Ala Glu Gly A Val Ala Glu Gly A
Val Ala Glu Gly G Val Ala Glu Gly G
Table 2. (Continued). (C) β-sheet and (D) turn/coil structure propensities, of amino acids in
the universal genetic code table. Letters in red, yellow and blue boxes represent large,
middle, and small β-sheet and turn/coil propensities, respectively. Meanings of color boxes
in Table (C) and (D) are the same as in Table (A) and (B), described above. Secondary
structure (β-sheet; (C) and turn/coil; (D)) propensities of amino acids were obtained from
Stryer’s “Biochemistry” (Berg et al, 2002)
2. Significance of the Genetic Code for life
The genetic code plays a quite important role in transfer of genetic information on DNA
nucleotide sequence to amino acid sequence of a protein, such as enzyme and transporter of
a chemical compound, etc (Figure 4). But, the genetic code has been generally regarded as a
simple representation of the relationship between a genetic information or a codon

transferred into mRNA is translated to the corresponding amino acid sequence of a protein
(Step 3) through genetic code mediating genetic information and catalytic function. The
universal genetic code used by extant organisms on the earth is composed of 64 codons and
20 amino acids (see Table 2)
3. Origin of the Genetic Code (GNC-SNS primitive genetic code hypothesis)
Our studies on the origin of the genetic code were initiated from the search for a prospective
spot on a DNA sequence, from which an entirely new gene encoding an entirely new
functional protein will be created, when an extant organism using the universal genetic code
has to adapt to a new environment. The spot was searched based on the six necessary
conditions for producing water-soluble globular proteins as described below. The six
conditions used for the search are hydropathy, α-helix, β-sheet and turn/coil formabilities,

Advances in the Study of Genetic Disorders
10
acidic amino acid and basic amino acid contents of proteins, which were obtained as
average values plus/minus standard deviations of water-soluble globular proteins in extant
micro-organisms. From the results, it was found that non-stop frames, which appear on anti-
sense strands of GC-rich genes (GC-NSF(a)s) at a high probability, have the strongest
possibility to create entirely new genes, not new modified type of genes or homologous
genes (Figure 5) (Ikehara et al., 1996). Where GC-NSF(a) means nonstop frame on antisense
strand of GC-rich gene. That is because hypothetical proteins encoded by GC-NSF(a)s
satisfied the six conditions and because the probability of non-stop frame (NSF) appearance
on the GC-rich anticodon sequences was enough high (Ikehara, 2002).
The GC-NSF(a) hypothesis on creation of the first family genes under the universal genetic
code led us propose subsequent theory on the origin of the genetic code as GNC-SNS
primitive genetic code hypothesis (Ikehara et al., 2002). GNC and SNS represent four
codons (GUC, GCC, GAC and GGC) and 16 codons (GUC, GCC, GAC, GGC, GUG, GCG,
GAG, GGG, CUG, CCG, CAG, CGG, CUC, CCC, CAC and CGC), respectively. I describe
the clues briefly below, from which the hypothesis was obtained. The first one is that base
sequences of the GC-NSF(a)s were rather similar to the repeating sequences of SNS. The

p
t
Maturation from a NSF(a) to a New GC-rich Gene
a GC-rich gene (an original gene)
a GC-rich gene a GC-rich gene
a GC-NSF(a)
a new GC-rich "original ancestor gene"

Origin of the Genetic Code and Genetic Disorder
11
was more primitive one than SNS by using the four more essential conditions which acidic
amino acid and basic amino acid compositions were excluded from the six conditions
described above. From the results, it was found that [GADV]-proteins encoded by GNC
codons well satisfied the four structural conditions, when roughly equal amounts of
[GADV]-amino acids were contained in the proteins (Figure 6 (B)). Where [GADV]
represents four amino acids of Gly, Ala, Asp and Val, and square bracket ([ ]) was used to
discriminate amino acids, especially G and A which are described by one-letter symbols of
amino acids, from nucleic acid bases, G and A. It means that even the [GADV]-polypeptide
chains with a quite simple amino acid composition could be folded into water-soluble
structures at a high probability.

(A) (B)

Fig. 6. (A) Dot plot analysis of SNS genetic code. Dots concentrated in the respective boxes
indicate that the six conditions (hydropathy, α-helix, β-sheet and turn/coil formabilities,
and acidic and basic amino acid contents) were satisfied. It means that polylpeptide chains
encoded by SNS code could be folded into water-soluble globular structures when bases are
contained in the respective rates at three codon positions. (B) Dot plot analysis of GNC code
On the other hand, other codes encoding four amino acids, which were picked out from the
columns or rows in the universal genetic code table, did not satisfy the four structural

n

(
%
) 100
0/100
0/100
0
50 50/100
50/100
100
100
50
GC Content (%)
50 60 70 80 90 100
100
100/0
100/0
100/0
0
50
50
50
50

Advances in the Study of Genetic Disorders
12
structures when four bases are contained in the respective rates at the second codon
position.
Thus, I provided GNC-SNS hypothesis as the origin of the genetic code about ten years ago
(Ikehara et al., 2002), suggesting that the universal genetic code originated from GNC code
through SNS code as capturing new codons up and down in the genetic code table (Figure 7
(B)).

(A) (B)

U C A G
Phe Ser Tyr Cys U
U Phe Ser Tyr Cys C
Leu Ser Term Term A
Leu Ser Term Trp G
Leu Pro His Arg U
C Leu Pro His Arg C
Leu Pro Gln Arg A
Leu Pro Gln Arg G
Ile Thr Asn Ser U
AIle ThrAsn Ser C
Ile Thr Lys Arg A
Met Thr Lys Arg G
Val Ala Asp Gly U
G Val Ala Asp Gly C
Val Ala Glu Gly A
Val Ala Glu Gly G
Fig. 7. GNC-SNS hypothesis on the origin and evolutionary pathway of the genetic code.
(A) In the hypothesis, it is supposed that the universal genetic code originated from GNC

Evolutionary process of the genetic code from GNC code, encoding four amino acids with
quite different chemical/physical properties, to the universal genetic code through SNS
code arranged amino acids with similar chemical and physical properties in the same
columns and with largely different properties in the same rows at high probabilities (Table
2). So, it is considered that the robustness of the genetic code originated from the
evolutionary process of the genetic code as suggested by the GNC-SNS primitive genetic
code hypothesis. The discussion on the robustness of the genetic code is consistent with the
results of permissible amino acid substitutions, which were observed between two
homologous proteins, as given in Figures 2 and 3. As described below, the finding of the
GNC-SNS primitive genetic code hypothesis led to the ideas on protein 0
th
-order structures
and on the origin of life as GADV hypothesis or [GADV]-protein world hypothesis (Ikehara,
2005; Ikehara, 2009).
4. The universal genetic code and protein 0
th
-order structure
Discussion on protein structure formation usually begins with primary structure or amino
acid sequence of a protein, not with amino acid composition. In Stryer’s textbook
“Biochemistry” (Berg et al, 2002), it is described that the information needed to specify the
catalytically active structure of ribonuclease is contained in its amino acid sequence. The
studies on folding of polypeptide chains, which were mainly carried out with small-sized
proteins, have established the generality of this central principle of biochemistry: sequence
specifies conformation. One of the reasons may rely on the facts that one-dimensional base
sequences on DNA or genes encode amino acid sequences or primary structure of proteins.
On the other hand, I happened to use amino acid composition for investigation of protein
structure formability, the six or four conditions as described above. The utilization gave
interesting results and conclusions, such as GC-NSF(a) hypothesis on creation of the first
family genes and GNC-SNS primitive genetic code hypothesis as described in the previous
Sections 3. During the investigation on the origin of the genetic code, I have noticed the