Description of sequence changes:  examples protein-level

Since references to WWW-sites are not yet acknowledged as citations, please mention den Dunnen JT and Antonarakis SE (2000). Hum.Mutat. 15:7-12 when referring to these pages.

Contents

• Introduction
  • reference sequence
• Examples  -  protein changes
  • general
  • six elementary content changes
    • substitutions  (missense, nonsense, nonstop)
    • deletion
    • duplication  (variability of short sequence repeats)
    • insertion 
    • inversion  (description not used at protein level)
    • translocation  (fusion proteins)
  • complex rearrangements
    • deletion/insertion (indel, frame shift)
  • miscellaneous
    • two changes in one allele
    • two change in one individual - allele unknown
    • changes in different alleles  (recessive diseases)
  • uncertainties  (exact position not known; Southern blot, PCR, arrayCGH, SNP-array, ...)
• Examples  -  DNA changes
• Examples  -  RNA changes

Introduction

Within this page examples will be given for the description of sequence variants on protein level, examples to describe changes at DNA and RNA level are given at other pages. All examples are described relative to a reference sequence, here the amino acid (protein) sequence. 

Reference sequence

Legend:
Reference sequence of imaginary gene used for the exaples given on this page. Nucleotide +1 in the coding DNA reference sequence is the A of the ATG translation initiation codon. Abbreviations used: nt = nucleotide, nts = nucleotides, UTR = untranslated region of the mRNA. For a picture of part of this hypothetical sequence see Figure.

General

It should be noted that the descriptions at protein level, even more than those at RNA level, are mostly deduced and not based on experimental evidence. Publications describing changes at protein level should make it clear whether experimental proof was available or not. In fact, when changes are reported for which experimental proof is not available one should consider to list them between brackets.

Sequence changes at protein level are basically described like those at the DNA level, with a few modifications;

• the three-letter amino acid code is prefered (see Discussion), with "X" designating a translation termination codon; for clarity we this page describes changes using the three-letter amino acid
• amino acid numbering
  • descriptions start with the amino acid, followed by its number (like p.Cys24 or p.C24)
  • the translation initiator Methionine is numbered as +1
  • amino acids after the stop codon (here after X110) are numbered relative to the stop codon as Gly110+1, Trp110+2, etc. 
• "silent" changes
  description of so called "silent" changes in the format p.Leu54Leu (or p.L54L) is not allowed; descriptions should be given at DNA level, it is non-informative and not unequivocal (there are five possibilities at DNA level which may underlie p.Leu54Leu);  correct description has the format c.162C>G.

Substitutions

Substitutions can be described without using the specific ">"-character which is used on DNA and RNA level (i.e. p.Trp26Cys in stead of p.Trp26>Cys).

• translation initiation site (Methionine codon, see Discussion)
  example: at DNA ATG (Met) changes to CTG (Leu) - description depends on the consequences of the change on the translation product (protein) produced;
  • effect unknown (most cases) - p.0?  (alternatively p.Met1?)
  • no protein produced - p.0  (experimental data should be available)
  • new upstream translation initiation site - p.[Met1extMet-8; Met1Leu] (alternatively p.[M1extM-8; M1L])
    NOTE: effectively this is an insertion (of Met-8 to Thr-1 between Thr-1 and Met1) combined with an amino acid change (Met1Leu)
  • new downstream translation initiation site - p.Met1_His15del (alternatively p.M1_H15del)
    NOTE: effectively this is a deletion 
• one amino acid to
  • another amino acid (missense change)
    Tryptophan 26 to a Cysteine; p.Trp26Cys (alternatively p.W26C)
  • a stop codon (nonsense change)
    Tryptophan 26 to a stop codon; p.Trp26X (alternatively p.W26X)
• translation termination site (stop codon, nonstop change) 
  • p.X110SerextX*17 (alternatively p.X110SextX*17)
    the stop codon (X) at position 110 is changed to a codon for Glutamine (Gln, Q), adding a tail of 16 new amino acids to the protein's C-terminus after which a new stop codon is reached
    NOTE: effectively this is a substitution (X110Ser) combined with an insertion (from Gly*1 to the new stop codon X*17)

NOTE: polymorphic variants are sometimes described as p.36Leu/Ile (p.36L/I) or p.36Leu/Leu (p.36L/L) but this is not correct (see Discussion).

Deletions

Deletions are designated by "del" after a description of the deleted segment, i.e. the first (and last) amino acid(s) deleted.

• MKLGHQQQCC to M_LGHQQQCC is described as p.Lys2del (alternatively p.K2del)
• MKLGHQQQCC to MKL___QQCC  is described as p.Gly4_Gln6del (alternatively p.G4_Q6del)
• MKLGHQQQCC to MKLGHQQCC is described as p.Glndel (p.Q8del)
  NOTE: for deletions in single amino acid stretches or tandem repeats, the most 3' residue is arbitrarily assigned to have been deleted
• if a deletion creates a new amino acid at the deletion junction the change is described as a insertion/deletion (see indels)

Duplications

Duplications are designated by "dup" after a description of the duplicated segment, i.e. the first (and last) amino acid(s) duplicated.

• MKLGHQQQCC to MKLGHQGHQQQCC is described as p.Gly4_Gln6dup (alternatively p.G4_Q6dup)
• MKLGHQQQCC to MKLGHQQQQCC is described as p.Gln8dup (alternatively p.Q8dup)
  NOTE: for duplications in single amino acid stretches or tandem repeats, the most 3' residue is arbitrarily assigned to have been duplicated
• MKLGHQQQCC to MKLGHQHQQQCC is described as p.His5_Gln6dup (alternatively p.H5_Q6dup)
  NOTE: duplicating insertions in single amino acid stretches (or short tandem repeats) should be described as a duplication and not as an insertion - so in the example shown p.Gln6_Gln7insHisGln (alternatively p.Q6_C7insHQ) is not correct
• variability of short sequence repeats are designated as p.Gln6(3_6) (alternatively p.Q6(3_6)) describing that the Glutamine (Gln, Q) stretch starting at position 6 in MKLGHQQQCC is found repeated 3 to 6 times in the population

Insertions

Insertions are designated by "ins" after a description of the amino acids flanking the insertion site, followed by a description of the inserted amino acids. When the insertion is large it may be described by its length (e.g. p.K2_L3ins34) but the inserted sequence should be described in detail in a footnote or as the accession number of the sequence as submitted to a sequence database (Genbank, EMBL, DDJB). Duplicating insertions should be described as duplications (see Discussion).

• MKLGHQQQCC to MKQSLGHQQQCC is described as p.Lys2_Leu3insGlnSer (alternatively p.K2_L3insQS)
• insertion of a 345 nucleotide sequence in intron 3; the sequence of the insertion need to be submitted to a sequence database (Genbank, EMBL, DDJB) and the accession numbers should be given

When an insertion creates a new amino acid at the insertion junction the change is described as an insertion/deletion (see indels)

Translocations

Translocations at protein level occur when a translocation at DNA level leads to the production of a fusion protein, joining the N-terminal end of the protein on one chromosome to the C-terminal end of the protein on the other chromosome (and vice versa). No recommendations have been made sofar to describe protein translocations.

• t(X;17)(DMD:p.Met1_Val1506; SGCA:p.Val250_X387) describes a fusion protein resulting from a translocation between the chromosomes X and 17; the fusion protein contains an N-terminal segment of DMD (dystrophin, amino acids Methionine-1 to Valine-1506), and a C-terminal segment of SGCA (alpha-sarcoglycan, amino acids Valine-250 to the stop codon at 387)

Complex rearrangements

Complex rearrangements are rearrangements which consist of several different types of the six elementary content changes substitution, deletion, duplication, insertion, inversion and translocation. Such rearrangements can be very complex and difficult to describe. Specific recommendations to describe such changes have not made. Complex rearrangements can be best described as a combination of the elementary changes. 

Deletion/insertions (indels) are described as a deletion followed by an insertion (see Discussion)

• p.Cys28_Lys29delinsTrp (alternatively p.C28_K29delinsW) denotes a 3 bp deletion affecting the codons for Cysteine-28 and Lysine-29, substituting them for a codon for Tryptophan
• p.Cys28delinsTrpVal (alternatively p.C28delinsWV) denotes a 3 bp insertion in the codon for Cysteine-28, generating codons for Tryptophan (W) and Valine (V)
• frame shift changes (see Discussion) cause, from a specific point onwards, the replacement of the normal C-terminal end of a protein for a new segment, encoded by the shifted reading frame. Frame shift changes can thus be best considered as deletion/insertions (indels). Frame shifts are designated by "fs" after the amino acid(s) affected by the change. Descriptions either use a short ("fs" only) or long ("fsX#") notation; the long description should include the change occurring at the site of the frame shift (see Discussion). In "fsX#", "X#" indicates at which codon position the new reading frame ends in a stop codon (X). The position of the stop in the new reading frame is calculated starting at the first amino acid that is changed by the frame shift, and ending at the first stop codon (X#). Thus, effectively, on DNA/RNA level the change might be one or more coding triplets up- or downstream.
  NOTE: the shifted reading frame is thus open for '#-1' amino acids.
  • p.Arg97ProfsX23 (short p.Arg97fs) denotes a frame shifting change with Arginine-97 as the first affected amino acid, changing into a Proline and the new reading frame ending in a stop at position 23
  • p.Arg97HisfsX5 (short p.Arg97fs) denotes a frame shifting change deleting amino acids Arginine-97 to Tryptophan-99, changing into a Histidine at the deletion junction and ending in the new reading frame in a stop at position 5
  • p.Leu30SerfsX3 (short p.Leu30fs) denotes a frame shifting change that deletes amino acids Leucine-30 to Cysteine-42 (exon 3 of the gene), substituting these for a Serine at the deletion junction and ending in the new reading frame in a stop at position 3

Miscellaneous

• Two changes in one allele are described as "[first change; second change]" (see Discussion)
  • p.[Trp13X; Pro43Ala] (alternatively p.[W13X; P43A]) describes a Tryptophan-13 to stop codon and Proline-43 to Alanine change in one allele (on one chromosome)
• Two changes in one individual when unknown in which allele are described as "[first change (+) second change]" (see Discussion)
  • p.[Trp13X(+)Glu61Gln] (alternatively p.[W13X(+)E61Q]) describes a Tryptophan-13 to stop codon and Glutamine-61 to Glutamic acid change in one individual while it is unknown whether these changes are on the same or different alleles
• Recessive disease (one change in each allele) are are described as "[change allele-1]+[change allele-2]" (see Discussion)
  • p.[Trp13X]+[Cys28Arg] (alternatively p.[W13X]+[C28R]) describes a Tryptophan-13 to stop codon change in one allele (chromosome) and a Cysteine-28 to Arginine change in the other allele (chromosome)
  • p.[Trp13X]+[?] (alternatively p.[W13X]+[?]) describes a Tryptophan-13 to stop codon change in one allele (chromosome) and an unknown change in the other allele (chromosome)
  • p.[Trp13X]+[=] (alternatively p.[W13X]+[=]) describes a Tryptophan-13 to stop codon change in one allele (chromosome) and a normal sequence in the other allele (chromosome)
  • two changes in different genes  -  SGCA:p.[Arg175X]+SGCB:p.[Cys305Ser] - the SGCA gene contains a nonsense change, the SGCB gene a Cys-to-Ser substitution
• When the effect on protein level is unknown, changes can be best described as "p.?" (see also above)

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