ANNUAL REPORT - Department of Biotechnology

At Birla Institute for Scientific Research, Jaipur,
studies have been carried out for development of
thermostable nitrile metabolizing enzymes for
enantio- and regio-selective biotransformation.
Streptomyces sp. MTCC 7546 was identified as the
best micro organism with desired characteristics and
the enzyme production from this strain is optimized.
The enantio-selectivity using mandelonitrile as
substrate in different percentage of organic solvent
was thoroughly investigated and an ee% of more
than 98% was achieved. The immobilization of
bacteria in agar-agar powder as an entrapment was
attempted and it was observed that the immobilized
whole cells could be used for more than 25 cycles
without much decrease in activity.
At NCL, Pune, biochemical and structural
investigations of pharmaceutically important
enzymes are being pursued. Fermentation
parameters for maximum production of
cephalosporin acylase from a new bacterial source,
Alcaligenes xylosoxidans ATCC 14648, have been
optimised. Chemical modification studies of the
purified enzyme revealed that serine plays an
important role in catalysis of the enzyme. Studies on
the substrate specificity of the enzyme indicated that
alpha amino substitution does not affect binding but
prevents catalysis. Crystal structure of gamma
Glutaryl trans peptidase from Bacillus subtilis has
been determined.
Drug Development and Delivery Systems
In a DBT funded project, KEM Hospital, Mumbai in
collaboration with Delhi University had developed a
liposomal amphotericin (Fungisome), which has
been investigated in animals and man, and shown to
be safe and effective. The manufactured formulation
is patented and currently available in the Indian
market. A multi centric, prospective, open label,
randomized trial comparing fungisome (1mg/kg/day
or 3 mg/kg/day) and conventional amphotericin B is
being conducted to investigate the use of liposomal
amphotericin in empirical therapy for presumed
fungal infection in febrile neutropenic patients. 6 out
of 7 patients enrolled in the study are assessable.
Three patients are randomized to the dose of
1mg/kg/day (Fungisome), and all are assessable.
Similarly 02 patients are randomized to 3 mg/kg/day
(Fungisome) dose, of which 01 is assessable. Two
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patients randomized to 1mg/kg/day (conventional)
are assessable.
In another multicentric, open, comparative,
randomized study to optimize dose, duration, safety,
efficacy, and cost of two treatment regimens with
Liposomal amphotericin (Fungisome) in the
treatment of systemic fungal infections in India, with a
sample size of 60, a total of 26 patients has been
enrolled so far at various centers of which 20 are
assessable. The analytical method for the
estimation of the Amphotericin B has been
developed and the validation is under progress.
At AIIMS, New Delhi, a mouse monoclonal antibody
against a neutralizing epitope of hepatitis B surface
antigen was cloned and expressed as mouse scFv
fusion protein of mouse scFv with human Fc
chimaeric Fab (fusion protein of mouse variable) and
human constant regions and full length chimaeric
antibody incorporating mouse variable and human
constant regions. All these molecules bind to the
same epitope on HbS Ag as the original monoclonal
and with similar affinity. A humanized antibody has
been expressed and characterized by further
mutating framework residues within the variable
regions of the mouse derived sequences to residues
found in human antibodies. These amino acids have
been identified by both homology matching and
molecular modeling.
In a multidisciplinary collaborative project at NII,
JNU, AIIMS and Kirorimal College, Delhi, with an aim
to inhibit beta cell autoimmunity in children
predisposed to get type I diabetes, the investigators
have designed competitive peptides to auto-antigens
in order to inhibit their presentation and abrogate
self-destruction of beta cells. The newly developed
altered peptides reduce the number of Th1 cells in
response to auto antigens when used for priming the
lymphocytes before exposure to the auto antigen invitro.
For in-vivo as well as targeted and sustained
delivery, these peptides have been encapsulated in
nanosized carriers. The release kinetics of the
peptides in buffer as well serum/plasma have been
studied. Two peptides with good results were
synthesized using standard FMOC chemistry and
purified using HPLC. Additional blood samples from
type 1 diabetes were studied to check if they would
inhibit auto-antigen-specific Th1 responses in